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44 Cards in this Set
- Front
- Back
What has been the biggest challange of producing the human plasma proteome? |
Retaining the structural integrity of proteins whilst extracting each and every one |
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10^ liters of what is PicoLitres? What is the volume of an average human cell? What is the average ATP concentration of an average cell? |
10^-12L 2pL 10mM |
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How many genes are present in the human genome? Of these, how many are needed? |
25,000 genes in total, about 18,000 needed |
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Roughly, how many proteinsare found in the human body? Roughly, how many metabolites are present? How many forms of sugar are present? |
1,000,000 proteins 3,000 metabolites 10,000 sugars |
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Of the cell homogenisation techniques, state 3 non-mechanical ones? |
Answer can include any 3 of the following: -Osmotic shock -Freeze thaw -Lytic enzymes -Lysozymes |
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Of the cell homogenisation techniques, state 5 mechanical ones? |
Answer can include any 5 of the following: -pestle and mortar+abrasives -Ball mills and glass beads -Blenders and rotor stators -Homogenisers -Solid extrusion -Liquid extrusion -Ultra-sonication |
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Osmotic Shock: What type of cell (s) would it be suitable for? State a disadvantage of using this techniques. |
Animal soft tissues, some plant cells Small scale only |
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Freeze/thaw: What type of cell (s) would it be suitable for? State a disadvantage of using this techniques. |
Animal soft tissues, some bacteria Answer can include: Time consuming, small scale, some enzymes cold-liable |
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Lytic enzymes: What type of cell (s) would it be suitable for? State a disadvantage of using this techniques. |
Animal cells and plant cells Answer can include: Small scale, expensive, enzymes must be removed once lysis is complete |
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Lysozyme: What type of cell (s) would it be suitable for? State a disadvantage of using this techniques. |
Some bacteria Gram negative must be treated with EDTA |
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Pestle, motar and abrasives: What type of cell (s) would it be suitable for? State a disadvantage of using this techniques. |
Tough tissues Not suitable for delicate tissues |
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Ball mills and glass beads: What type of cell (s) would it be suitable for? State a disadvantage of using this techniques. |
Bacteria and fungi May cause organelle damage in eukaryotes |
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Blenders and rotor stators: What type of cell (s) would it be suitable for? State a disadvantage of using this techniques. |
Plant and animal tissues Ineffective for microbes |
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Homogenisers (glass and Teflon): What type of cell (s) would it be suitable for? State a disadvantage of using this techniques. |
Soft, delicate tissues Glass may shatter-goggles needed |
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Solid extrusion (Hughes Press): What type of cell (s) would it be suitable for? State a disadvantage of using this techniques. |
Tough plant material, bacteria and yeast Small scale |
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Liquid Extrusion (French Pressure cell): What type of cell (s) would it be suitable for? State a disadvantage of using this techniques. |
Microbial cells Small scale |
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Ultrasonication: What type of cell (s) would it be suitable for? State a disadvantage of using this techniques. |
Microbial cells Answer may include: cooling required, small scale, may cause damage to organelles, especially in eukaryotic cells |
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There are 8 components needed in a disruption media. State 6 of these. |
>Buffer-replace internal buffer,usually pH 7 >Inorganic salts-KCl and NaCl, usually below >100mM >Sucrose-prevents osmotic lysis of organelles >Mg2+ ions-improve integrity of membranes >EDTA >Protease inhibitors >Reducing agents >Detergents |
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State the purpose of having: EDTA Protease inhibitors Reducing agents Detergents In a Disruption Media? |
>EDTA: chelate heavy metal contaminations, remove ionic Ca2+ >Protease inhibitors: Prevent breakdown of proteins (use PMSF) >Reducing agents: Prevent oxidation of proteins by disulfide bridges, sulfenic and sulfinic acids >Detergents: cause dissociation of proteins and lipoproteins from the cell membrane |
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State the 5 properties of proteins that can be measured. |
Mass Charge-pH Hydrophobic/hydrophilic properties Differential solubility Mobility in applied fields |
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To isolate a protein from a cell, conditions within the cell have to be replicated. What 5 things need to be controlled? What also does it need protection from? What also needs to be preserved? |
pH, ionic strength, viscosity, solvation and temperature Proteases Maintain oxidation state |
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There are 8 non-polar hydrophobic amino acids. State 5 of them. |
Answer can include any 5 of the following: Alanine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan and valine |
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There are 7 hydrophilic, un charged, polar, amino acids. State 5 of them. |
Answer can include any 5 of the following: Asparagine, cysteine, glutamine, glycine, serine, threonine and tyrosine |
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There are 5 charged, polar amino acids. State 2 positively charged ones and 2 negatively charged ones. |
Positively charged: (Answer can include)Arginine, histidine and lysine Negatively charged: Aspartic acid and glutamic acid |
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Give the formal definition of pH as an equation. |
pH=-Log(alphaH3O+) |
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In the above equation, what does alpha H3O+ represent? |
Activity of the hydrated hydrogen ion |
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Give the definition of Activity, as an equation. |
Activity=gamma[H3O+] |
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What does gamma represent in the above equation? |
Gamma: activity co-efficient, giving a value of between 0-1 |
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Define equilibrium constant as an equation. |
Ka=[H3O+][A-]/[AH] |
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Express the answer to the above flashcard logarithmically. |
pKa=-Log10Ka |
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A lower pKa produces a higher Ka. What kind of acid is this? |
Stronger acid |
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pKa of an acid is the pH when...? |
It is exactly half dissociated |
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pH greater than the pKa, the acid is? What about when the pH is less than the pKa? |
Acid is fully dissociated Acid is predominantly HA |
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The following equation may be derived from which equation (give its name): pH=pKa+Log10([A-]/[HA]) |
Henderson-Hasselbach Equation |
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When [A-]=[HA] what does pH=? |
pKa |
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Potential difference is generated between 2 electrodes immersed in solution. State the equation used to calculate this. (Draw this out on paper) |
V=E^o+2.303RT/FxpH |
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In the above equation, what does: V, E^o, R, T and F signify? |
V: measured voltage E^o: reference potential of the Ag/AgCl R: gas constant J K^-1 M^-1 T: temperature K F: Faraday's constant CM^-1 |
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Define the isoelectric point. |
pH at which the molecule contains no net electric charge |
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Define a buffer. |
Resists a change in H+ concentration (pH) on addition of acid or alkali |
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What identifies a good buffer on a Buffer Titration Curve? |
A flat plateau |
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There are 6 properties to an ideal buffer. State 4 of them. |
Answer can include any 4 of the following: >Impermeable to biological membranes >Biological stability and lack of interferences with metabolic and biological processes >Lack of significant absorption of ultraviolet or visible UV light >Lack of formation of insoluble complexes with cations >Minimal effect of ionic composition or salt concentration >Limited pH change in response to temperature |
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Give an example of a Zwitterionic buffer. |
HEPES |
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Seperation of proteins. State 2 ways that mass can separate proteins. State 2 ways that affinity can separate proteins. |
Centrifusion Passage through membranes (dialysis) Hydrophobic/hydrophilic property columns Differential solubility extraction buffers |
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State 3 ways that Charge and Mobility in Applied Fields can be used to seperate proteins. |
Buffer control Separation on Gels Electrophoresis |