Bms1027 Proteins

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Amino acids

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nh2 amine group attached to a central carbon which is also bonded to a h, cooh and r group.
can be essential (cannot be synthesised internally) or non essential (synthesised by converting other amino acids into one another)

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Primary structure

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- condensation reaction of amino acids with the release of h2o
- joined by peptide bonds

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Secondary structure

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Hydrogen bonds form to create alpha helix's and beta pleated sheets out of polypeptide chain

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tertiary structure

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folding of protein change due to hydrophobic interactions and the creation of electrostatic and disulfide bonds. Can be globular or fibrous.

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Quaternary structure

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2 or more tertiary structures join to form a globular or fibrous complexes

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Conjugated proteins

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Protein complexes with metal or prosthetic groups bonded to them
eg lipoproteins, glycoproteins, metalloproteins, nucleoproteins, phophoproteins

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Derived proteins

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proteins treated by a chemical or enzymes
eg alkaline extraction of soya protein or rennin during cheese manufacture

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R groups (types)

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- polar, can hydrogen bond with water (soluble)
- non polar, insoluble in water
- Positively charged, have extra amino group
- negatively charged, have extra COOH group
- sulfide, can form disulfide bonds

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Chemical reactions - COOH group

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- esterification
- decarboxylation
- reduction to OH

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Chemical reactions - amine group

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- acylation with acid anhydride to give acyl amino acids (modifies properties - may make essential amino acids unavailable to body)

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Chemical reactions - disulfide group (S-S)

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- splitting forming disulfide bonds (bread making)
- thermal gelation (in non reversible gels/foams)
- discolouration in canned low acid, high protein foods (disadvantage)

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Chemical reactions - sulphydryl groups (S-H)

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- oxidise free radicals from oxidative rancidity
- active sites for enzymes
- bind metals
- protein cross linking in S-H/S-S interchange

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Chemical reactions - Nitrite groups

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- added during curing to give flavour, colour and protection against spores
- can form nitrosamines (toxic - disadvantage)

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Chemical reactions - Maillard browning

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- reaction between the amino group of a protein and the carbonyl group of a sugar
- gives some desirable products (odour, colour, taste etc) but also possibly toxic products and makes essential amino acids unavailable

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Chemical reactions - Formation of lysinoalanine

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- under alkaline conditions amino acids cystine and serine break down to dehydroalanine which reacts with lysine to form lysinoalanine
- product isnt digestible as peptide bond form cant be broken down, also possibly toxicity and makes essential amino acids unavailable

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Denaturation

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- intermediate 3`/4` bonds break causing unfolding
- caused by: heat, freezing, pH, chemicals, physical conditions (pressure, shear)
- can be reversible if mild conditions, but extremes form irreversible denaturations to form aggregates
- aggregates form when hydrophobic group rearrange onto outside of protein causing them to clump together and ppt out

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Enzymes

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- biological catalysts that speed up reactions without being altered by the process
- have active site which forms weak interactive bonds with a substrate to form a complex (induced fit)
- have optimum conditions: pH (affects ionisation so alters ionic bonds between as and s), temp (denature after certain temp, to low and there low activity so slow rate)

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Common food enzymes

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- ISOMERASE: converts glucose to fructose which is sweeter so less is needed in food
- INVERTASE: converts sucrose to glucose and fructose to produce an invert sugar for confectionary
- LIPOXYGENASE: converts unsat FA to FA hyperoxides used during bread making to improve dough
- PROTEASE: used to tenderise meat and during cheese manufacture
- LACTASE: breaks lactose into galactose and glucose, can be intolerant so need supplement or dairy foods treated with lactase

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Analysis - Application

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- nutritional value determination/labelling
- protein purification
- legal requirements
- functional, structural, enzymatic properties
- product development

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Analysis - Total nitrogen determination (kjeldahl method)

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- principle: digest sample in conc H2SO4, and then in NaOH, steam distil product and collect in inert boric acid. Titrate against 0.1M HCL to work out total N2
- catalyst: kjeldahl tablets contain mercuric oxide or selenium to shorten digestion time and potassium/sodium sulphate to raise temp
- ADVAN: reference method, direct results, reliable
- DISADVAN: measures non-protein N2 also, not very sensitive (gram quantities required)

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Analysis - Indirect colourmetic methods (biuret)

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- principle: reaction of protein with dye to get purple colour, the intensity can be measured using a spectrophotometer at 540nm against a calibration curve. dye reacts with peptide bonds
- ADVAN: simple, quick, doesnt depend on AA type (all peptide bonds are the same)
- DISADVAN: not very sensitive

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Analysis - Indirect colourmetric methods (lowry)

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- principle: react with Cu2+ under alkaline conditions, complex forms with folin reagent to give a blue product.
- ADVAN: very sensitive, widely used, simple
- DISADVAN: interference of lipids, sucrose, buffers etc, variation between proteins as reliant on tyrosine/tryptophan, pH and temp dependent

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Analysis - Dye binding assays

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- principle: use -ve dye to bind to +ve amino acids under acidic conditions which produces a ppt which can be centrifuged and the amount of protein calculated (absorbance against protein conc)
- ADVAN: very simple, sensitive, less interference from lipids, buffers etc
- DISADVAN: relies on +ve AAs, can have non-ionic hydrophobic interactions

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Analysis - Amino acid analysis

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- principle: acid hydrolysis in 8M HCL at 110c, separation by column chromatography (ion exchange column) to identify and detect AA's
- uses: nutritional quality (essential AA amount), flavour quality

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Analysis - HPLC

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- high pressure liquid chromatography
- ADVAN: fast and sensitive, use high pressure and small columns to produce a peak trace which is easy to analyse
- DISADVAN: need standard trace to compare to

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Functional properties - Protein

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- organoleptic: colour, texture, flavour, odour
- hydration: solubility, viscosity
- rheology: aggregation, elasticity, viscosity
- surface: emulsification, foaming

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Functional properties - Soya proteins

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- application: used in meat, dairy and bakery products
- water binding: has good binding potential
- gelation: isolates form pasty gels (need high conc)
- texturisation: can be extruded under high pressure/temp to form meat substitutes and fibres
- emulsification: can bind fat
- foaming: has poor foaming properties

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Functional properties - Egg proteins

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- white: ovalbumin (heat gelation), conalbumin (gelation), ovomucoid (inhibits trypsin, foaming)
- yolk: LDL (aeration, stabilisation), HDL and livetins (emulsification)
- storage: deterioration caused by microbes, to avoid can store in low temps, coat with sodium silicate to seal pores, control humidity and ^ CO2. Deterioration causes thinning of white, reduction of Mg and Ca, weakening of vitelline membrane, evaporation of water, increase in air sac size.

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Functional properties - Flour and bread proteins (1)

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- Milling: wheat is cleaned and conditoned before milling, bran and germ can be removed and endosperm ground into powder
- Protein displacement milling: separates flour into fractions by air classification depending on starch granule size, and thus protein conc. (allows range of flours from one parent flour)

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Functional properties - Flour and bread proteins (2)

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- if just endosperm ground then white flour produced, can have bran or germ added back, needs added chalk to increase Ca content and vit B/iron additives
- wholegrain flour has nothing removed or added but does contain phytic acid from bran which forms insoluble complexes with Ca and Fe making them unavailable

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Functional properties - Dough making/baking

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- Conventional: single process, dough fermented at 30c, proofed to provide CO2 for leavening and modification of dough proteins, baking at 230c
- Mechanical: batch or continuous, bulk fermentation replaces by intense short time mixing, chemical oxidation with ascorbic acid, can also add 0.7% fat for gas retention and 3.5% extra water. (stales slower, increases yield, uses weak uk flour and quicker)
- Microwave: uniform cooking, reduces amylase activity which is important in starch degradation so need high amylase flour. can use low protein flour as gases produced faster than lost

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Functional properties - Flour improvers

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- ascorbic acid or cystine
- act on SS and SH bond

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Functional properties - Gluten

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- Gliadin (43%), small globular proteins, soluble in ethanol and extensible
- Glutenins (39%) larger fibrous proteins, suspendible in solvents, complexes with lipids and low extensibility
- some other proteins, fats, sugars etc make up the rest of gluten
- gluten makes up 85% of flour proteins

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Functional properties - Non-gluten proteins

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- soluble so washed away, makes up 15% flour proteins
- eg, Albumins and Globulins (contain useful enzymes and coagulation proteins)

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Functional properties - Milk proteins (whey)

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- 0.6% of milk
- small globulin proteins inc beta lactoglobulin, bovine serum albumin
- denatures easily and can produces gels for desserts, has good foaming properties and a good emulsifier (can act as a egg substitute)

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Functional properties - Milk proteins (casein)

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- exists as spherical micelles, made of 2 types, alpha, beta, kappa and gamma
- sub micelles consist of hydrophobic core, surrounded by kappa casein shell with calcium phosphate's on surface which adhere sub micelles together
- stable of heating, isoelectric point of 4.6, good emulsifier, hydrolysed to peptides for flavours, foams and emulsions, bioactive peptide (neutraceuticals)

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Functional properties - Milk proteins (products)

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- pasteurised = kills bacteria only, 72c for 15s
- sterilised = kills bacteria and spores, 80c for 20-40s
- in can sterilised = 130c for 3-5 mins
- UHT = 149c for 1sec plus aseptic processing and packaging

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Functional properties - Milk proteins (cheese)

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- process: 1) coagulation with rennin or acid 2) whey drained away and moisture removed by development of acid, continued rennin action, heating, stirring or pressing 3) curd shaped and cut into pieces 4) curd cured to improve flavour, texture etc.
- Rennin action = attacks specific bond in k-casein which causes it to ppt out and clot in the presence of Ca, which results in a fibrous network
- RENNET CURD = elastic, hard (40-50% moisture), high fat as envelopes fat and insoluble salts
- ACID CURD = fragile, non-elastic, gelatinous, insoluble salts are soluble in whey, low fat, 70% moisture

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Functional properties - Milk proteins (ice cream)

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- process: mixing, pasteurisation (80c for 10min), homogenisation, cool to 4c ageing (2-3hrs), freezing and whipping
- types: hard, soft or whippy scoop
- stabilisers: carrageenan prevents separation, carob gum increases water holding capacity and heat shock properties, perin also a low pH to be maintained in fruit ice cream

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Functional properties - Meat proteins (muscle structure)

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- Myofibrillar proteins: MYOSIN (large charged molecule with strong affinity to Ca2+/Mg+ ions, 2 sub units (light and heavy meromyosin), HMM contains ATPase and bonded to actin during contraction. Two myosin units twist around each other to form a dimer) ACTIN (smaller, globular units combines to form helical filament, associated with troponin, actinin & tropomyosin)
- Connective tissue: COLLAGEN (insoluble in salt and water, produces gelatin on heating/hydrolysed, triple helix structure with high hydroxyproline/lysine content) RECTICULIN (doesnt produce gelatin) ELASTIN (ligaments, not broken by heating)

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Functional properties - Meat proteins (muscle to meat)

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1) oxygen level falls after death
2) respiration ceases
3) anaerobic respiration (glycolysis) starts, producing lactic acid and causing the Ph to fall
4) energy rich phosphate bonds break (ATP, GP etc) and levels diminish
5) onset of rigor mortis (muscle fibres fixed)
6) proteins denature
7) enzymes released causing fat oxidation etc, accumulates metabolites and aids flavour development

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Functional properties - Meat proteins (effect of glycogen)

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- low glycogen causes low levels of lactic acid formation so a higher ultimate pH of 6 producing a dark glazy meat
high glycogen causes high levels of lactic acid formations so a lower ultimate pH of 5 and PSE meat (pale, soft and exudative)

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Functional properties - Meat proteins (conditioning)

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- meat conditioned by holding it at 1c for a few days to cause actin to break away from Z lines which softens and tenderises meat to improve flavour (can cause bacterial spoilage, denaturation and desiccation)
- DENATURATION: changed of 2`/3` structures, loss of solubility and biological activity. Myofibrillar proteins denature easily and are affected by temp, produces a high ultimate pH.
- tenderisation depends on muscle contraction at on set of rigor mortis, if extended then little actomyosin and tender meat, if contracted then much cross linking so tough meat.

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Functional properties - Meat proteins (nutrition)

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- good source of essential amino acids and fatty acids (can be saturated = atheroschlorosis/CVD etc)
- lots of minerals (Fe, K, Ph etc)
- lots of vitamins (B1, B2, A, folic acid)

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Functional properties - Meat proteins (properties)

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1) colour: depends on quantity/state of myoglobin (whether its oxidised or not), temp of cooking, diffusion of O2. Discolouration occurs due to changes in ultimate pH, bacteria and yellow fat
2) water holding capacity: depends on ultimate pH, rigor mortis, conditioning, freezing
3) texture: preslaughter (species, age, type of muscle, conditon/treatment before death) post slaughter (rigor mortis, ultimate pH etc) conditioning and processing (freezing, artificial tenderisation , cooking)
4) odour/taste: chemical aspects (water/fat soluble precursors, volatiles, amino acids+sugars can be used to create artificial meat flavours

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Functional properties - Meat proteins (processing)

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1) chilling: carcass chilled quickly in air controled room to avoid spoilage and drying out. PVC used for packaging to allow O2 diffusion to maintain colour whilst in fridge
2) freezing: rapid freezing with prior chilling can led to loss of fluid on thawing. Storage can cause oxidation of fats so needs very low temp and O2 impermeable packaging opaque/shrink wrap/edible gaze also helps prevent drying and spoilage)
3) thermal processing: pasteurisation, sterilisation
4) moisture control: by direct removal (sun/oven drying), or by increased extracellular osmotic pressure by the addition of brine (curing) - smoking optional after curing (uses smoke liquid phase and dispersed gas phase of acids/phenols/hydrocarbons etc)