BioInorganic Chemistry of Zinc Chemistry 3391B - Instruct
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BioInorganic Chemistry of Zinc
Chemistry 3391B
*
Instructor: Martin Stillman ChB064
Martin.stillman@uwo.ca
Executive summary:
A boring metal: *d10 – no colour – zinc proteins are colourless unlike Fe & Cu proteins.
Amazing biological chemistry. Remarkable versatility.
Chem 3391B :ZINC R20-jklM
3391b-20202-D-ZINC-R20jklML.docx 2020-03-01 6:03:00 PM
Page 1 of 32 Chem 3391B:ZINC R-20-M
The role of zinc (an essential group 12 metal) 1. Zinc is everywhere in our environment 2. Zinc is a constituent of over 3000 enzymes involved in numerous body functions, including enzymes involved in gene expression, wound healing, growth. 3. Deficiency impairs cell growth and repair of tissue injury; leads to poor growth, delayed wound healing, impairment of sexual development and decreased taste acuity. 4. Zinc is present in gustin, a salivary polypeptide that is necessary for the development of taste buds. 5. Its binding motifs are: Intermediate-Intermediate or S-I or H-I (ligand-metal order) – such a versatile metal! 6. HIS and CYS are favourite amino acids – plus water and carboxylic acids (GLU, ASP) in the 3rd and 4th position if chemistry takes place there. TETRAHEDRAL geometry – sometimes 5 groups but from 4 different ligands – ie the carboxylic group of GLU or when acting as a protease. Proteins of interest: 1) Metallothionein (7 Zn) 4 CYS; -below 2) Carbonic anhydrase (CA) 3 HIS 1 water; - below 3) 2) Carboxypeptidase A (CPA) 2 HIS; 1 GLU; 1 water; - below 4) 3) Alcohol dehydrogenase (ADH) (2 Zn) 2 CYS; 1 HIS; 1 water; - below 5) 5) Zn – finger proteins typically - CYS-CYS HIS-HIS (C2H2 motif) - briefly below (other motifs are known: 4CYS; - note donor atoms: CYS – SH HIS =N- or –NH-; ASP & GLU -OH & =O) Page 2 of 32 Chem 3391B:ZINC R-20-M
Let’s return to our chart from the first lectures:
About 2 g in a 70 g human – right up next to Fe. •Good sources meat, liver, eggs and seafood are good
RDA: for adults - males: 15 mg & females: 12 mg dietary sources, whereas zinc in vegetable sources
(green leafy vegetables and legumes), and particularly
Zinc Requirement and Effects of Deficiency
in cereal grains, is less bioavailable.
Zinc is essential in -
Beef is a major source of iron and zinc for American
1. Development and functioning of the brain.
children and adults – actually is the number one
2. Zinc dependent enzymes and neurotransmitters source of zinc
play a role in the central nervous system, which
may in turn affect cognition. Supplement: no more than 40 mg (above this is
considered toxic)
3. Important for healthy skin, a healthy immune
system, and resistance to infection.
Pumpkin seeds – 1/2 cup: 8.4 milligrams (>50% RDA!)
4. Wound healing with Zn ointments (from the ancient
1 veggie hot dog 3.75 mg
world).
1 cup cooked greens 0.8 mg
5. Growth is dependent on Zn. Emerging research on ¼ cup peanuts……… 3.6 mg
human models supports a beneficial role for Zn in ¼ cup pumpkin seeds… 2.6 mg
human cognitive development and function 1 cup Cheerios……… 4.5 mg
6.
Emerging research on human models supports a
beneficial role for zinc in human cognitive
development and functioning
Page 3 of 32 Chem 3391B:ZINC R-20-M
Page 4 of 32 Chem 3391B:ZINC R-20-M
http://www.healthaliciousness.com/articles/zinc.php 12.3mg (82% 38.7mg (258%
14.2mg (95% DV)
DV) DV)
Lamb is also a good source of Zinc (%DV per 3oz cooked): Lean
Top 10 Foods Highest in Zinc Foreshank (49%), Lean Shoulder (46%) and Lean Cubed Lamb for
Stewing (37%). Click to see complete nutrition facts.
Zinc is an essential mineral required by the body for maintaining a
sense of smell, keeping a healthy immune system, building proteins, #3: Wheat Germ (Toasted)
triggering enzymes, and creating DNA. Zinc also helps the cells in
your body communicate by functioning as a neurotransmitter. A
Zinc in 100g Per Cup (113g) Per Ounce (28g)
deficiency in zinc can lead to stunted growth, diarrhea, impotence,
hair loss, eye and skin lesions, impaired appetite, and depressed 16.7mg (111% DV) 18.8mg (126% DV) 4.7mg (31% DV)
immunity. Conversely, consuming too much zinc can lead to nausea,
vomiting, loss of appetite, abdominal cramps, diarrhea, and Crude or Untoasted Wheat Germ is also a good source of Zinc
headaches in the short term, and can disrupt absorption of copper providing 94% DV per cup:. )
and iron in the long term. If you have a zinc deficiency, then animal
foods are better sources of zinc than plant foods. The current
daily value (DV) for Zinc is 15mg. Below is a list of the top ten
foods highest in Zinc by common serving size, for more, see #4: Spinach
the list of high zinc foods by nutrient density, and the extended
list of zinc rich foods.
Zinc in 100g Per Cup (Cooked -
100g (Raw)
(Cooked) 180g)
#1: Seafood (Cooked Oysters) 0.5mg (4%
0.8mg (5% DV) 1.4mg (9% DV)
DV)
Per 6 Oysters Other Green Leafy Vegetables High in Zinc (%DV per cup):Amaranth
Zinc in 100g Per 3oz (85g) Leaves, cooked (8%), and Endive and Radiccio, raw (2%). Click to see
(42g)
complete nutrition facts.
78.6mg (524% 66.8mg (445%
33.0mg (220% DV)
DV) DV) #5: Pumpkin and Squash Seeds
Other Seafood High in Zinc (%DV per 3oz cooked): Crab (43%), and
Lobster (41%). Click to see complete nutrition facts.
Zinc in 100g Per Cup (64g) Per Ounce (28g)
#2: Beef and Lamb (Cooked Lean Beef Short ribs) 10.3mg (69% DV) 6.6mg (44% DV) 2.9mg (19% DV)
Other Seeds High in Zinc (%DV per ounce): Sunflower (10%), Chia
(9%), and Flaxseeds (8%). Click to see complete nutrition facts.
1 Rack of Ribs 1 Lean Ribeye Fillet
Zinc in 100g
(315g) (129g)
Page 5 of 32 Chem 3391B:ZINC R-20-M
#6: Nuts (Cashews) #10: Mushrooms (Cooked White Mushrooms)
Zinc in 100g Per Ounce Per Cup Pieces Per Mushroom
Per Cup (137g) Zinc in 100g
(Roasted) (28g) (156g) (12g)
7.7mg (51% 0.9mg (6%
5.6mg (37% DV) 1.6mg (10% DV) 1.4mg (9% DV) 0.1mg (1% DV)
DV) DV)
Other Nuts High in Zinc (%DV per ounce): Pine nuts (12%), Pecans Other Mushrooms High in Zinc (%DV per Cup Pieces): Morel, raw
(9%), Almonds (6%), Walnuts (6%), Peanuts (6%), and Hazelnuts (5%). (9%), Brown, raw and Portabella, grilled (5%), Oyster, raw (4%), and
White, raw (2%). Four Dried Shitake mushrooms contain 8% DV and 4
raw shitake contain 4% DV.
#7: Cocoa and Chocolate (Cocoa Powder) Recipes High in Zinc
(Click to expand)
Zinc in 100g Per Cup (86g) Per Tablespoon (5g) Buckwheat (How to Cook Buckwheat)
6.8mg (45% DV) 5.9mg (39% DV) 0.3mg (2% DV) Teff (How to Cook Teff)
Dark baking Chocolate is also high in Zinc providing 85% DV per Blackberry Salad
cup grated and 19% DV per 29g square. Click to see complete TOP TEN by Zn density per serving
nutrition facts.
78.6mg (524% DV) 66.8mg (445% DV) per
#8: Pork & Chicken (Cooked Lean Pork Shoulder) #1: Oysters (Cooked)
per 100 grams 3oz (85 grams)
#2: Wheat Germ 16.7mg (111% DV) 4.7mg (31% DV) per
Zinc in 100g Per Steak (147g) Per 3oz (85g)
(Toasted) per 100 grams ounce (28 grams)
5.0mg (33% DV) 7.4mg (49% DV) 4.3mg (28% DV)
Chicken is also High in Zinc providing 15% DV per cooked 12.3mg (82% DV) 38.7mg (258% DV) per
#3: Beef (Lean, Cooked)
drumstick. Click to see complete nutrition facts. per 100 grams piece (315 grams)
11.9mg (79% DV) 8.0mg (53% DV) per
#9: Beans (Cooked Mung Beans) #4: Veal Liver (Cooked)
per 100 grams slice (67 grams)
Zinc in 100g Per Cup (124g) Per 3oz (85g) #5: Pumpkin & Squash 10.3mg (69% DV) 2.9mg (19% DV) per
Seeds (Roasted) per 100 grams ounce (28 grams)
0.5mg (3% DV) 0.6mg (4% DV) 0.4mg (2.5% DV)
Other Beans High in Zinc (%DV per cup cooked): Baked Beans Note this web site - invaluable in reporting nutrition per food
(39%), Adzuki (27%), Chickpeas (17%) and Kidney Beans (12%). Click http://nutritiondata.self.com/foods-000124000000000000000.html
to see complete nutrition facts.
Page 6 of 32 Chem 3391B:ZINC R-20-M
Very useful data from an NIH web - note - several within the context of a total diet. The DV for zinc is 15 mg for adults and
children age 4 and older. Food labels, however, are not required to list zinc
pages content unless a food has been fortified with this nutrient. Foods providing
http://ods.od.nih.gov/factsheets/Zinc- 20% or more of the DV are considered to be high sources of a nutrient.
The U.S. Department of Agriculture's (USDA's) Nutrient Database Web site
HealthProfessional/ [11] lists the nutrient content of many foods and provides a comprehensive list
Table 2: Selected Food Sources of Zinc [11] of foods containing zinc arranged bynutrient content and by food name.
Milligrams
(mg) Percent Summary of benefits - much more below:
Food per serving DV*
Oysters, cooked, breaded and fried, 3 ounces 74.0 493 Healthy Immune Function - Even mild to moderate zinc deficiency can
Beef chuck roast, braised, 3 ounces 7.0 47 depress the immune system through impaired macrophage and
Crab, Alaska king, cooked, 3 ounces 6.5 43 neutrophil functions, and associated effects.3 Zinc is also essential for
Beef patty, broiled, 3 ounces 5.3 35 creation and activation of T-lymphocytes.4,5 Further, low levels of zinc
Breakfast cereal, fortified with 25% of the DV for 3.8 25
have been associated with increased susceptibility to pneumonia and
zinc, ¾ cup serving
other infections in children and the elderly.6-9
Lobster, cooked, 3 ounces 3.4 23
Pork chop, loin, cooked, 3 ounces 2.9 19 Alleviation of the Common Cold (*Controversial) - There are
Baked beans, canned, plain or vegetarian, ½ cup 2.9 19 conflicting studies as to weather or not zinc supplements can alleviate
Chicken, dark meat, cooked, 3 ounces 2.4 16 symptoms of the common cold and shorten its duration. At least one
Yogurt, fruit, low fat, 8 ounces 1.7 11 study confirms decreased duration of cold symptoms compared to a
Cashews, dry roasted, 1 ounce 1.6 11 control,10 however, other studies report no effect.11,12 Since no harm is
Chickpeas, cooked, ½ cup 1.3 9
reported, increasing zinc intake could only help.
Cheese, Swiss, 1 ounce 1.2 8
Healing of Cuts and Wounds - Zinc is essential for healthy skin and
Oatmeal, instant, plain, prepared with water, 1 1.1 7
maintenance of mucosal membranes. Adequate levels of zinc is
packet
Milk, low-fat or non fat, 1 cup 1.0 7 necessary for proper wound healing.13
Almonds, dry roasted, 1 ounce 0.9 6 Reduced Severity and Duration of Diarrhea - Studies show that
Kidney beans, cooked, ½ cup 0.9 6 increased intake of zinc can reduce duration and severity of diarrhea in
Chicken breast, roasted, skin removed, ½ breast 0.9 6 undernourished children with infections.14-17
Cheese, cheddar or mozzarella, 1 ounce 0.9 6 Prevention and Reduction of Age-Related Eye Damage - High dietary
Peas, green, frozen, cooked, ½ cup 0.5 3
intake of zinc, as well as vitamins C, E, and beta-carotene, has been
Flounder or sole, cooked, 3 ounces 0.3 2
associated with reduced age-related macular deneration in the edlerly.
* DV = Daily Value. DVs were developed by the U.S. Food and Drug
Administration to help consumers compare the nutrient contents of products
Page 7 of 32 Chem 3391B:ZINC R-20-M
------not required to learn the text from here to p 11 taking zinc supplements [61]. The children in these studies received 4–40 mg
of zinc a day in the form of zinc acetate, zinc gluconate, or zinc sulfate [61].
– for general interest In addition, results from a pooled analysis of randomized controlled trials of
Zinc deficiency is characterized by growth retardation, loss of appetite, and zinc supplementation in developing countries suggest that zinc helps reduce
impaired immune function. In more severe cases, zinc deficiency causes hair
the duration and severity of diarrhea in zinc-deficient or otherwise
loss, diarrhea, delayed sexual maturation, impotence, hypogonadism in males,
and eye and skin lesions [2,8,24,25]. Weight loss, delayed healing of wounds, malnourished children [62]. Similar findings were reported in a meta-analysis
taste abnormalities, and mental lethargy can also occur [5,8,26-30]. Many of published in 2008 and a 2007 review of zinc supplementation for preventing
these symptoms are non-specific and often associated with other health and treating diarrhea [63,64]. The effects of zinc supplementation on diarrhea
conditions; therefore, a medical examination is necessary to ascertain
in children with adequate zinc status, such as most children in the United
whether a zinc deficiency is present. Taste from gustin.
States, are not clear.
Immune function
Severe zinc deficiency depresses immune function [48], and even mild to The World Health Organization and UNICEF now recommend short-term zinc
moderate degrees of zinc deficiency can impair macrophage and neutrophil supplementation (20 mg of zinc per day, or 10 mg for infants under 6 months,
functions, natural killer cell activity, and complement activity [49]. The body for 10–14 days) to treat acute childhood diarrhea [60].
requires zinc to develop and activate T-lymphocytes [2,50]. Individuals with The common cold
low zinc levels have shown reduced lymphocyte proliferation response to Researchers have hypothesized that zinc could reduce the severity and
mitogens and other adverse alterations in immunity that can be corrected by duration of cold symptoms by directly inhibiting rhinovirus binding and
zinc supplementation [49,51]. These alterations in immune function might replication in the nasal mucosa and suppressing inflammation [65,66].
explain why low zinc status has been associated with increased susceptibility Although studies examining the effect of zinc treatment on cold symptoms
to pneumonia and other infections in children in developing countries and the have had somewhat conflicting results, overall zinc appears to be beneficial
elderly [52-55]. under certain circumstances. Several studies are described below in which zinc
Wound healing is administered as a lozenge or zinc-containing syrup that temporarily "sticks"
Zinc helps maintain the integrity of skin and mucosal membranes [49]. in the mouth and throat. This allows zinc to make contact with the rhinovirus
Patients with chronic leg ulcers have abnormal zinc metabolism and low serum in those areas.
zinc levels [56], and clinicians frequently treat skin ulcers with zinc In a randomized, double-blind, placebo-controlled clinical trial, 50 subjects
supplements [57]. The authors of a systematic review concluded that zinc (within 24 hours of developing the common cold) took a zinc acetate lozenge
sulfate might be effective for treating leg ulcers in some patients who have (13.3 mg zinc) or placebo every 2–3 wakeful hours. Compared with placebo,
low serum zinc levels [58,59]. However, research has not shown that the the zinc lozenges significantly reduced the duration of cold symptoms (cough,
general use of zinc sulfate in patients with chronic leg ulcers or arterial or nasal discharge, and muscle aches) [67].
venous ulcers is effective [58,59]. In another clinical trial involving 273 participants with experimentally induced
Diarrhea colds, zinc gluconate lozenges (providing 13.3 mg zinc) significantly reduced
Acute diarrhea is associated with high rates of mortality among children in the duration of illness compared with placebo but had no effect on symptom
developing countries [60]. Zinc deficiency causes alterations in immune severity [68]. However, treatment with zinc acetate lozenges (providing 5 or
response that probably contribute to increased susceptibility to infections, 11.5 mg zinc) had no effect on either cold duration or severity. Neither zinc
such as those that cause diarrhea, especially in children [49]. gluconate nor zinc acetate lozenges affected the duration or severity of cold
symptoms in 281 subjects with natural (not experimentally induced) colds in
Studies show that poor, malnourished children in India, Africa, South America, another trial [68].
and Southeast Asia experience shorter courses of infectious diarrhea after
Page 8 of 32 Chem 3391B:ZINC R-20-M
In 77 participants with natural colds, a combination of zinc gluconate nasal associated with high zinc intakes. After an average follow-up period of 6.3 spray and zinc orotate lozenges (37 mg zinc every 2–3 wakeful hours) was also years, supplementation with antioxidants plus zinc (but not antioxidants found to have no effect on the number of asymptomatic patients after 7 days alone) significantly reduced the risk of developing advanced AMD and reduced of treatment [69]. visual acuity loss. Zinc supplementation alone significantly reduced the risk of In September of 2007, Caruso and colleagues published a structured review of developing advanced AMD in subjects at higher risk but not in the total study the effects of zinc lozenges, nasal sprays, and nasal gels on the common cold population. Visual acuity loss was not significantly affected by zinc [66]. Of the 14 randomized, placebo-controlled studies included, 7 (5 using supplementation alone. A follow-up AREDS2 study confirmed the value of this zinc lozenges, 2 using a nasal gel) showed that the zinc treatment had a supplement in reducing the progression of AMD over a median follow-up period beneficial effect and 7 (5 using zinc lozenges, 1 using a nasal spray, and 1 of 5 years [75]. Importantly, AREDS2 revealed that a formulation providing 25 using lozenges and a nasal spray) showed no effect. mg zinc (about one-third the amount in the original AREDS formulation) More recently, a Cochrane review concluded that "zinc (lozenges or syrup) is provided the same protective effect against developing advanced AMD. beneficial in reducing the duration and severity of the common cold in healthy Two other small clinical trials evaluated the effects of supplementation with people, when taken within 24 hours of onset of symptoms" [70]. The author of 200 mg zinc sulfate (providing 45 mg zinc) for 2 years in subjects with drusen another review completed in 2004 also concluded that zinc can reduce the or macular degeneration. Zinc supplementation significantly reduced visual duration and severity of cold symptoms [65]. However, more research is acuity loss in one of the studies [76] but had no effect in the other [77]. needed to determine the optimal dosage, zinc formulation and duration of A Cochrane review concluded that the evidence supporting the use of treatment before a general recommendation for zinc in the treatment of the antioxidant vitamins and zinc for AMD comes primarily from the AREDS study common cold can be made [70]. [71]. Individuals who have or are developing AMD should talk to their health As previously noted, the safety of intranasal zinc has been called into question care provider about taking a zinc-containing AREDS supplement. because of numerous reports of anosmia (loss of smell), in some cases long- Interactions with iron and copper lasting or permanent, from the use of zinc-containing nasal gels or sprays [14- Iron-deficiency anemia is a serious world-wide public health problem. Iron 16]. fortification programs have been credited with improving the iron status of Age-related macular degeneration millions of women, infants, and children. Fortification of foods with iron does Researchers have suggested that both zinc and antioxidants delay the not significantly affect zinc absorption. However, large amounts of progression of age-related macular degeneration (AMD) and vision loss, supplemental iron (greater than 25 mg) might decrease zinc absorption [2,78]. possibly by preventing cellular damage in the retina [71,72]. In a population- Taking iron supplements between meals helps decrease its effect on zinc based cohort study in the Netherlands, high dietary intake of zinc as well as absorption [78]. beta carotene, vitamin C, and vitamin E was associated with reduced risk of High zinc intakes can inhibit copper absorption, sometimes producing copper AMD in elderly subjects [73]. However, the authors of a systematic review and deficiency and associated anemia [79,80]. For this reason, dietary supplement meta-analysis published in 2007 concluded that zinc is not effective for the formulations containing high levels of zinc, such as the one used in the AREDS primary prevention of early AMD [74], although zinc might reduce the risk of study [72], sometimes contain copper. progression to advanced AMD. The Age-Related Eye Disease Study (AREDS), a large, randomized, placebo- controlled, clinical trial (n = 3,597), evaluated the effect of high doses of Health Risks from Excessive Zinc selected antioxidants (500 mg vitamin C, 400 IU vitamin E, and 15 mg beta- Zinc toxicity can occur in both acute and chronic forms. Acute adverse effects carotene) with or without zinc (80 mg as zinc oxide) on the development of of high zinc intake include nausea, vomiting, loss of appetite, abdominal advanced AMD in older individuals with varying degrees of AMD [72]. cramps, diarrhea, and headaches [2]. One case report cited severe nausea and Participants also received 2 mg copper to prevent the copper deficiency Page 9 of 32 Chem 3391B:ZINC R-20-M
vomiting within 30 minutes of ingesting 4 g of zinc gluconate (570 mg Penicillamine
elemental zinc) [81]. Intakes of 150–450 mg of zinc per day have been Zinc can reduce the absorption and action of penicillamine, a drug used to
associated with such chronic effects as low copper status, altered iron treat rheumatoid arthritis [87]. To minimize this interaction, individuals
function, reduced immune function, and reduced levels of high-density should take zinc supplements at least 2 hours before or after taking
lipoproteins [82]. Reductions in a copper-containing enzyme, a marker of penicillamine [85].
copper status, have been reported with even moderately high zinc intakes of Diuretics
approximately 60 mg/day for up to 10 weeks [2]. The doses of zinc used in the Thiazide diuretics such as chlorthalidone (Hygroton®) and hydrochlorothiazide
AREDS study (80 mg per day of zinc in the form of zinc oxide for 6.3 years, on (Esidrix® and HydroDIURIL®) increase urinary zinc excretion by as much as 60%
average) have been associated with a significant increase in hospitalizations [88]. Prolonged use of thiazide diuretics could deplete zinc tissue levels, so
for genitourinary causes, raising the possibility that chronically high intakes of clinicians should monitor zinc status in patients taking these medications.
zinc adversely affect some aspects of urinary physiology [83].
The FNB has established ULs for zinc (Table 3). Long-term intakes above the
UL increase the risk of adverse health effects [2]. The ULs do not apply to http://www.berkeleywellness.com/supplements/miner
individuals receiving zinc for medical treatment, but such individuals should
als/article/zinc-fighting-cold-war
be under the care of a physician who monitors them for adverse health
effects. thumbs-up
Table 3: Tolerable Upper Intake Levels (ULs) for Zinc [2]
In 2011, zinc made headlines again, thanks to a review from the
Age Male Female Pregnant Lactating
Cochrane Collaboration, which evaluates scientific research. The
0–6 months 4 mg 4 mg
review, which was updated in 2013, concluded that, compared to a
7–12 months 5 mg 5 mg
placebo, zinc can shorten colds by about a day and reduce their
1–3 years 7 mg 7 mg
severity somewhat, particularly when started within 24 hours of the
4–8 years 12 mg 12 mg
first symptoms. (Another 2012 research review on zinc, published
9–13 years 23 mg 23 mg
in theCanadian Medical Association Journal, came to similar
14–18 years 34 mg 34 mg 34 mg 34 mg
conclusions, though it noted that “uncertainty remained about its
19+ years 40 mg 40 mg 40 mg 40 mg
clinical benefit.”)
Interactions with Medications
Zinc supplements have the potential to interact with several types of
The Cochrane research is not new research, but is an analysis of
medications. A few examples are provided below. Individuals taking these data from 15 “good-quality” studies done since 1987. The studies
medications on a regular basis should discuss their zinc intakes with their included 1,360 healthy people who took lozenges, tablets or syrup
healthcare providers. in the form of zinc gluconate, acetate or sulfate, usually at the first
Antibiotics sign of a cold. Doses ranged from 30 to 190 milligrams a day. Not
Both quinolone antibiotics (such as Cipro®) and tetracycline antibiotics (such all the studies found a benefit. The main adverse effects were
as Achromycin® and Sumycin®) interact with zinc in the gastrointestinal tract,
nausea and a bad taste in the mouth.
inhibiting the absorption of both zinc and the antibiotic [84,85]. Taking the
In two of the studies in the review, children who took lower-dose
antibiotic at least 2 hours before or 4–6 hours after taking a zinc supplement
minimizes this interaction [86].
zinc syrup for five to seven months developed fewer colds. This
Page 10 of 32 Chem 3391B:ZINC R-20-Mwas the only evidence of a preventive effect, but these studies were
of poorer quality.
think about zinc
Like many nutrients, zinc has its good-guy/bad-guy aspects. It is
needed for cell division, growth, wound healing and the proper
functioning of the immune system, and also plays a role in acuity of
taste and smell.
Zinc is plentiful in some foods, such as meat and seafood. The
recommended dietary allowance, or RDA, for zinc is eight to 11
milligrams a day for people 14 or older, and three to eight
milligrams for children, depending on their age. Prolonged use of
high doses of zinc can interfere with the body’s absorption
of copper, as well as depress the immune system, impair blood cell
formation, and have other potential adverse effects. In adults, even
50 to 75 milligrams of zinc a day for long periods can be a problem,
and it would take even less to adversely affect children.
Bottom line: If you want to try zinc lozenges when you have a
cold, start as soon as you have symptoms and follow the dosing
advice on the label (usually every two hours), though admittedly
this is a shot in the dark. Don’t take the lozenges for more than a
week, and don’t take them long-term to prevent colds.
Page 11 of 32 Chem 3391B:ZINC R-20-MOK back to the course content….
Meat is much better as a source of Zn than cereals. High Zn is
found in the fetus – developmental reasons? 60
Zinc % of Food Supply
50 41.5
Mainly transported by the protein metallothionein (see below), 40
which controls absorption 30
18.9 18.4
20
Zinc deficiency can lead to: 10
8.6 6.3 6.3
1. Hair loss,
0
Meat Grain Legumes
Poultry Nuts
2. Delayed wound healing Fish Soy
3. Dwarfism when there is major zinc deficiency due to poor diet Food Groups
– was the case in rural Iran in 1960’s
4. Structural malformations in the brain
5. Changes in enzymes and proteins important for neurotransmission
6. Behavioural problems - reduced attention
7. Reduced memory - reduced ability to learn
8. Low zinc status during pregnancy may affect the development of the infant’s
nervous system and later cognitive functioning
9. In older adults (aged 65-90) dietary zinc is positively linked to cognitive
function
10. Low levels of zinc in the blood and brain tissues of patients with Alzheimer’s
Severe zinc deficiency is seen primarily in alcoholics (especially if they have
developed cirrhosis), patients with chronic renal disease or severe Zheng JJ, Mason JB, Rosenberg IH, et al.
malabsorption diseases Measurement of zinc bioavailability from beef and
a ready-to-eat breakfast cereal in humans:
application of a whole-gut lavage technique. Am J
Clin Nutr. 1993; 58: 902-907.
Page 12 of 32 Chem 3391B:ZINC R-20-M1999 11 12 13 14 15 16
Where are we in the
Periodic Table?
Group 12 (recent
numbering scheme for
groups)
All 2+ cations (well, Hg
can be 1+: [Hg2]2+)
As 2+ cations, all are d10
Become ‘softer’ down the
triad
Need to memorize this
triad
What is special about Zn(II)?
3d10 – so no contribution to bonding – bonds form between s and p orbitals – empty now.
No redox chemistry possible – it’s Zn(II) all the time. So why Zn(II)? That’s the point – it can
help make changes in organic molecules without being involved. It acts as a tether. Most
important changes the acid-base properties of ligands so that acid-base chemistry takes place
that shouldn’t at pH 7 – see CA mechanism below.
L-B R-M K-S Problems to do
1-2; 9 2 4 See the questions at
the end
Page 13 of 32 Chem 3391B:ZINC R-20-MZinc containing proteins and enzymes:
1) Metallothionein (MT)- 7 Zn (and Cd) 4 CYS ligands each metal –
tetrahedral coordination solely by CYS
7 METALS – 20 CYS
2 domains some CYS are shared – see (b) and (c) for the structures of the
metal binding sites.
Major zinc binding protein – acts as a storage site. Essentially no free zinc in
a cell. KF = 7 = ok - careful equals K1xK2… each = approx. 1011 ! So for 7
Zn(II) = a very large number….
(Sutherland.. et al. Biochemistry, 2012; Sutherland & Stillman, Metallomics, 2014)
Page 14 of 32 Chem 3391B:ZINC R-20-MIn enzymes, very many examples all Zn(II) dependent: 1) carbonic anhydrase (CA) 2) matrix metalloproteases (MMPs), secreted by cells – involved in degrade old tissue during tissue growth 3) carboxypeptidase A (CPA) 4) alcohol dehydrogenase (ADH) – from liver = LADH 5) zinc finger DNA-binding-proteins (and A1) 6) Superoxide dismutase (ZnCuSOD) – only at the end. 7) Phospholipase C structure - 3-Zn-enzymes ZINC in catalytic roles binds readily to HIS, CYS, GLU, ASP, and water. Zn2+ acts as a Lewis Acid – so, is an electron acceptor – binds anions – so the ligand usually becomes charged – the effect is a reduction over several orders of magnitude of the pKa of the group (makes the group dramatically more acidic) – BUT HAS TO HAVE A VACANT SITE – well a site that can accept the substrate – so occupied by water or OH- not a strong sigma donoer like N or S.. As an enzyme – major role is activating water (or ROH or R2C=O) – making the water (and other groups) far more acidic – that is changing H2O almost into OH-. BUT Binds to 4 cys – or 2 cys & 2 his or 4 his etc in structural roles Page 15 of 32 Chem 3391B:ZINC R-20-M
1) Carbonic anhydrase - CA Zn2+ activation of H2O in carbonic anhydrase Carbonic Anhydrase was the first enzyme identified that contained a zinc atom In CA, Zn2+ is bound to the enzyme through 3 Histidines + water – see the next pages for more details about the reaction = NOT a structural role- The overall reaction catalysed: How does it do this? And what is special about the zinc coordination site? It’s all to do with the histidine/imidazole nitrogen Intermediate ligand on an Intermediate metal – not a soft ligand like??___________). See next pages. Page 16 of 32 Chem 3391B:ZINC R-20-M
H
H H CO2 H+ O
O
Zn His O - O
His94 119
His96 His94 Zn His119
HCO3- H2O His96
H+ CO2 H2O HCO3-
L3 Zn-(OH2) -
PZn-(OH ) PZn-(HCO3-) PZn-(OH2)
Page 17 of 32 Chem 3391B:ZINC R-20-MMechanism of Carbonic Anhydrase
We can see the mechanism in more detail here:
Key – activation of the CO2 by the negatively charged OH on the zinc. Then the now
negatively charged O on the CO2 (the new Lewis base) binds to the zinc(II) – it’s the Lewis
acid. Now we have a Glu-like binding pattern.
Incoming water displaces the HCO3- product.
The H2O is activated by the Zn(II) – makes it more
acidic – so the H+ is lost at neutral pH rather than at
the much higher pH normally needed. The reaction
usually takes place at pH 7 – 8 – acidosis of the blood
would dramatically slow down the chemistry (above)
because there would be H+ present driving the
reaction to the LEFT.
CA – primarily a pleated structure
Page 18 of 32 Chem 3391B:ZINC R-20-M2) Zinc proteases (metalloproteases) include: digestive enzymes – eg carboxypeptidase A CP-A eg stem bromelain is a proteolytic enzyme from pineapple (does not contain metals) – in addition to your lips can tenderize meat The zinc-binding motif at the active site of a metalloprotease includes: 2 His residues whose imidazole side-chains are ligands & 1 Glu with 2 oxygens = catalysis NOT a structural role During catalysis, the Zn2+ promotes nucleophilic attack on the carbonyl carbon by the oxygen atom of a water molecule at the active site. The active site (Glu in carboxypeptidase) facilitates this reaction by extracting H+ from the attacking H2O. 3) Carboxypeptidase A – a pancreatic digestive enzyme that cleaves the carboxyl terminal amino acid from a peptide chain by hydrolysing the amide link: Example: LEU-GLU-PHE LEU-GLU + PHE This is the reverse of the peptide chain expansion reaction. Mass: 34,500, 1 Zn, the Zn is bound in a crevice with almost 5 coordination - HIS HIS GLU (both O’s) and water - catalysis NOT a structural role. To cleave, break up, digest the peptide chain, hence a peptidase. The organic chemistry says: nucleophilic attack of the C=O should work, but what with? Needs an OH- - where do we get that from? We need one bound to a Zn(II), but at pH 7 – not many OH- around, we have plenty of water though. The chemistry of the bound OH- is even more aggressive and has to be stabilized more. So, a 5-coordinate Zn(II) – really illegal – but possible with db metals because they can access the 4s, 4p and 4d (the 3d are filled) – sp3d This gives the OH- a great incentive to leave… which it does. Important to remember the following: Activate the water, make a stronger base (OH-) it attacks the C=O. Don’t forget the hydrogen bonds – really essential they keep the peptide held in place… Page 19 of 32 Chem 3391B:ZINC R-20-M
4) LIVER ALCOHOL DEHYDROGENASE LADH A 2-zinc-containing dimeric enzyme (=4 Zn), uses a coenzyme (NAD+) and ethanol. A zinc metalloenzyme that oxidizes alcohols to aldehydes or ketones – a member of the oxidoreductase family of enzymes – a redox enzyme – shuttles electrons. The catalytic zinc is bound to two CYS, 1 HIS and water; the 2nd zinc is bound to four CYS = a structural role.. In humans, the enzyme is contained in the lining of the stomach and in the liver. It catalyzes the oxidation of ethanol to acetaldehyde: CH3CH2OH + NAD+ → CH3CHO + NADH + H+ It is important to note that the aldehyde is more toxic than the alcohol and water is being used - so dehydration occurs – hence the hang-over – luckily aldehyde is broken down more rapidly… hmmmm This allows the consumption of alcoholic beverages, but its evolutionary purpose is probably the breakdown of alcohols naturally contained in foods or produced by bacteria in the digestive tract. Humans have at least six slightly different alcohol dehydrogenases. Serine (S48) provides the route for the proton, above. Page 20 of 32 Chem 3391B:ZINC R-20-M
Up to 30% of alcohol is oxidized in the stomach – class IV rxn. The rate-limiting step is the availability of the coenzyme, NAD+. We only have enough enzyme so that, for an average size person, it will take about 90 minutes to metabolize 1 drink (typical oxidation rate is 100 mg ethanol/kg/hr = 11 ml/hr/70 kg). The liver is the major site of this oxidation, as well as the site of major destruction (cirrhosis) due to excessive amounts of alcohol. The overall reaction catalyzed (Class I) is the oxidation of alcohol to aldehyde with the concomitant reduction of the coenzyme NAD+ to NADH: RCH2OH + NAD+ RCHO + NADH + H+ We can see how the enzyme works as follows: •Enz + NAD+ Enz-NAD+ •Enz-NAD+ + ETOH Enz-NAD+-ETOH •Enz-NAD+-ETOH Enz-NADH + aldehyde (RCHO) AND then •Enz-NADH Enz + NADH Ethanol metabolism in humans follows zero-order kinetics; unfortunately, not inducible (which means cant improve the rate rink). The rate-limiting step is the availability of the coenzyme, NAD+. `tep 4 – the release of NADH to cycle round to the 1st step. We only have enough enzyme such that, for an average size person, it will take about 90 minutes to metabolize 1 drink (typical oxidation rate is 100 mg ethanol/kg/hr = 11 ml/hr/70 kg individual). The liver is the major site of this oxidation, as well as the site of major destruction (cirrhosis) due to excessive amounts of alcohol. Page 21 of 32 Chem 3391B:ZINC R-20-M
SUMMARY 1 )Two identical subunits in LADH 2) The enzyme contains two Zn2+ per subunit; one is in the active-site and involved in catalysis, the other serves a structural role. For the active-site Zn the ligands are 2 Cys, 1 His and a water molecule. The structural Zn has 4 CYS ligands. 3) LADH is a major enzyme in alcohol metabolism, and is most concentrated in the liver. Since vitamin A is an excellent substrate it has been suggested that the "real" substrate is vitamin A – ie NOT alcohol - (the aldehyde of which is critically important in vision and epithelial tissue). 4) Ethanol metabolism in humans follows zero-order kinetics; unfortunately, not inducible (which means the more alcohol you drink …) 5) The rate-limiting step is the availability of the coenzyme, NAD+. 6) Step 4 – the release of NADH to cycle round to the 1st step. 7) We only have enough enzyme such that, for an average size person, it will take about 90 minutes to metabolize 1 drink (typical oxidation rate is 100 mg ethanol/kg/hr = 11 ml/hr/70 kg individual). 8) The liver is the major site of this oxidation, as well as the site of major destruction (cirrhosis) due to excessive amounts of alcohol. The catalytic chemistry involves protonation of the NAD+ the NAD+ is the required step – and controls the rate CH3CH2OH + NAD+ CH3CHO +NADH + H+- Page 22 of 32 Chem 3391B:ZINC R-20-M
5) …but what about the zinc fingers? CYS CYS HIS HIS – generally – Zn2+ 4 CYS or 2CYS and 2HIS for structural jobs – no empty or reactive sites left. Zinc-finger DNA binding protein family The DNA-binding motif is found as part of transcription regulatory proteins. Zinc finger proteins were first discovered as transcription factors. (SEE BELOW for details) Zinc finger proteins are among the most abundant proteins in eukaryotic genomes. Their functions are extraordinarily diverse include DNA recognition, RNA packaging, transcriptional activation, regulation of apoptosis, protein folding and assembly, and lipid binding. Zinc finger structures are as diverse as their functions. One of the most abundant DNA-binding motifs. Proteins may contain more than one finger in a single chain. Three common types: His2ZnCys2, Cys4Zn, or Cys6Zn2 Always TET. - see GAL-4 2Zn structure below: A web site to check out: http://labs.umassmed.edu/WolfeLab/Cys2His2_Zinc_Finger_Proteins.html Page 23 of 32 Chem 3391B:ZINC R-20-M
What do Zinc-Finger proteins do? Zn finger proteins are DNA-binding proteins They all exhibit the same motifs of 2 antiparallel beta sheets and a alpha helix linked together by the Zn. http://www.ebi.ac.uk/interpro/potm/2007_3/Page1.htm Protein of the Month The key is that the Zn holds the whole protein together and stabilizes the structure. Although several different ZnF (Zn-finger) motifs are known, the overall summary is that: ZnF may bind one or more Zn, have many fingers, and drape across DNA (and other hosts) by recognising in DNA nucleic acids with specific amino acid sequences (see below for examples). So, ZnF can locate themselves along a particular part of the DNA strands. The original and sometimes called 'classical' ZnF has the Zn motif of Cys2His2 - with a short beta hairpin and an alpha helix (beta/beta/alpha structure), where a single zinc atom is held in place by Cys(2)His(2) (C2H2), but also possible are Cys(2)HisCys (C2HC), or Cys(3)His (CCCH) residues. Multiple Zn's and therefore, multiple fingers are found. There is always the beta- beta- structures in each finger. Because each finger binds to 2 or 3 bases the recognition becomes more precise by reading more of the DNA sequence and the binding strength increases making binding more efficient. Page 24 of 32 Chem 3391B:ZINC R-20-M
Transcription factors use the Zn-finger on the DNA as a trigger. The -helices of the Zn-finger wrap around the DNA. John H Wilson Nature Biotechnology 21, 759 - 760 (2003) doi:10.1038/nbt0703-759 The middle finger from the Zif268 transcription factor illustrates the structure of a zinc-finger domain and identifies the amino acid residues important for DNA binding. The zinc-finger nuclease is modified from Figure 1 in Bibikova et al.10. Notice the amino acid residues that bind to the bases: They are? See the next page….for the 1-letter codes and then next, the larger diagram Page 25 of 32 Chem 3391B:ZINC R-20-M
Page 26 of 32 Chem 3391B:ZINC R-20-M
www.mrc-mbu.cam.ac.uk
Legend for figures: The canonical zinc finger
protein Zif268 recognizes its binding site using
three Cys2His2 zinc fingers [DNA recognition by
Zif268].
Each finger inserts its a helix into the major
groove, and DNA recognition is mediated by
amino acids at positions –1, 2, 3 and 6 from this
helix (numbered with respect to the start of the
alpha helix). The majority of contacts are made
to one strand of the DNA.
Interactions that are mediated by hydrogen
bonds between the fingers and DNA are
indicated by arrows in the panel on the right;
hydrophobic interactions are indicated by open
circles.
An alignment of the amino acid sequence of the
three Cys2His2 zinc fingers from Zif268 is shown
for reference. Adjacent fingers are connected by
‘canonical’ linkers (TGQKP and TGEKP). The
residues from each finger involved in defining
R=ARG; D=Asp (1 Ch2); E=GLU (2-Ch2); C=CYS; H=HIS DNA specificity are indicated in blue; residues
involved in zinc coordination are indicated in
purple.
The secondary structure elements are indicated
above the sequence with arrows indicating beta-
strands and the cylinder denoting alpha-helices.
Page 27 of 32 Chem 3391B:ZINC R-20-MSTRUCTURES - The following images are provided to allow us to examine how Nature uses the rules of inorganic chemistry to construct binding sites. Included to allow you to understand more completely the reactions and structures described above. Zinc binds here to 2 CYS and 2 HIS (2S and 2N). Fingers bind to 3 base-pair subsites and specific contacts are mediated by amino acids in positions -1, 2, 3 and 6 relative to the start of the alpha-helix. Contacts mainly involve one strand of the DNA. Where proteins contain multiple fingers, each finger binds to adjacent subsites within a larger DNA recognition site thus allowing a relatively simple motif to specifically bind to a wide range of DNA sequences. This means that the number and the type of zinc fingers dictate the specificity of binding to DNA Page 28 of 32 Chem 3391B:ZINC R-20-M
6) Superoxide dismutase (the binding of the Cu(I) and Zn(II) can be rationalised in terms of our models
in the inorganic unit and here) – SOD – implicated in ALS disease due to muscle and nerve degeneration. Note –
the O2- reaction was described in the inorganic unit. Here, one of a great many, CuZnSOD enzymes. Note –
needs catalase (Fe(III-IV)-heme enzyme) to decompose the H2O2 product.
2 O2- + 2H+ O2 + H2O2
The chemistry requires both metals. SOD activity first described
in a series of famous papers by Fridovich & McCord in 1969 The
link to Amyotrophic Lateral Sclerosis (als) was reported in 1993
Page 29 of 32 Chem 3391B:ZINC R-20-M7) Aminopeptidases - example AAP catalyzes cleavage of amino acid at the N-terminal position (whereas the carboxypeptidase cleaves from the C-terminal) A monomeric enzyme consisting of two Zn2+ ions; the active site consists of the two zinc centers bridged by Asp117 and an oxygen species obtained from a hydroxide group; two Zn are also coordinated with Glu151, Glu152, His97, His 256 and Asp179. Located in the small intestines to carry out the final stage of digestion of proteins to the constituent amino acids. Page 30 of 32 Chem 3391B:ZINC R-20-M
Key points from this unit
1 Relationship of Zn to other metals – what is its role? Why was the change from anaerobic
to aerobic atmosphere not significant for zinc?
Common binding site combinations for Zn – check out p 2 – what are the amino acids
involved?
What are the binding sites of Zn as a catalyst and zinc as a structural metal?
Why the difference?
What makes Zn such a good catalyst?
Why so much Zn do you think?
Zn deficiency is hard to achieve (is that a real sentence or thought?) but with not enough
Zn – problems occur like Pb excess (is Pb excess possible?)
So, Zn deficiency – cognitive problems – where is Zn deficiency really found? Liver
disease – from?
List, name , 3 Zn-containing enzymes – and 1 Zn structural-required protein
2 Metal distribution comes from natural and man-made sources – know the cycle- but is Zn
toxic? No!! Well, yes, in excess.
A major played in the transport of Zn is metallothionein – 7 Zn(II) per molecule, bound
only to CYS – we see this protein later as a major played for Cd metabolism and toxicity.
3 So, how does Zn act in enzymes? It is its Lewis acid properties – which are what?
Carbonic anhydrase – key is CO2 is polarised – OH- from water attacks CO2 – the key is
that H2O is made much wore acidic by the Zn. What are the other ligands in CA? Know
Page 31 of 32 Chem 3391B:ZINC R-20-Mthe cycle shown on p 16.
Carboxypeptidase – works the same way – know the key step of attack of C=O by O
attached to Zn. – see p. 17
Alcohol dehydrogenase – see p 24 – here the alcohol binds to the Zn –and the aldehyde is
formed – how does NAD+ enter the reaction? What is it called? Recognise its structure.
4 Zinc finger proteins bind to DNA – how? What does the Zn do?
Study questions from the lectures:
What is a Lewis acid? How does it differ from a protonic acid? Why is Zn called a Lewis acid? Name
a Lewis base. What is the key property of Zn that is exploited in the zinc- enzymes? How?
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