Biochem: AA, peptides and proteins Flashcards
How many AA are there?
20
Structure of AA
carboxyl group and an amino group bonded to the same carbon atom (the alpha carbon)
what are the non polar aliphatic AAs?
glycine (G), Leucin (L) Isoleucine (I), Methionine (M), Valine(V), Proline (P), Alanine (A)
Aromatic AAs
Phenylalanine (F), Tyrosine (Y), Tryptophan (W)
Polar, uncharged AAs
Serine (S), Threonine (T), Cysteine (C), asparagine (N), Glutamine (Q)
Positively charged AAs
Lysine (K), Arginine (R), Histidine (H)
Negatively charged AAs
Aspartate (D) , Glutamate (E)
Where would one usually find alanine and leucine in a folded protein?
in the core of the protein. Alanine and leucine are non polar, hydrophobic AAs. the hydrophobic side chains are attracted to the core of the protein in to avoid interaction with the aqueous environment, the hydrophobic nature of the AA stabilise the protein core via hydrophobic interactions.
What is the effect of the aliphatic side chain of proline on polypeptides containing it?
Proline has an aliphatic side chain with a distinctive cyclic structure. The secondary
amino (imino) group of proline residues is held in a rigid conformation that reduces the structural flexibility of polypeptide regions containing proline.
Protein X absorbs light at a wavelength of 280-300 nm at its active zone. Which AA is likely part of this region?
Tyrosine. Tyrosine and tryptophan (and to a lesser extent phenylalanine) are aromatic AAs that have the ability to absorb UV light. The hydroxyl group of tyrosine can form hydrogen bonds, and it is an important functional group in some enzymes.
Why does the titration curve of glycine have 2 stages?
Because it has 2 ionisable chains. When glycine is titrated, both the carboxyl and amino group are ionised using a strong acid like NAOH. The resulting two stages in the plot correspond to deprotonation of the two different groups on glycine
How can you determine if an AA is charged?
By titration. The net electric charge of an AA is reflected at its isoelectric point. in the example of glycine, since it does not have a charged side chain, the pI is the mean between the carboxyl and amino groups’ pKa (2.97+9.6/2 = 5.97) –> since it does not have a charged R group, at pH = 5.97 glycine is in its zwitterion form, and thus uncharged. At any pH below its pI, glycine has a net positive charge and will move toward the negative electrode (the cathode)
What is a peptide bond?
condensation reaction between 2 AAs
a protein only contains AA residues and no other chemical groups. how can we know the number of AA in the protein?
divide the total weight by 110
What is the most powerful way to separate (fraction) proteins?
column chromatography
How does column chromatography work?
It takes into account differences in size, charge and polarity of proteins.
- A porous solid material with appropriate chemical properties (the stationary phase) is held in a column, and a buffered solution
(the mobile phase) migrates through it. - The protein, dissolved in the same buffered solution that was used to establish the mobile phase, is layered on the top of the
column. - The protein then percolates through the solid matrix as an ever-expanding band within the larger mobile phase.
- Individual proteins migrate faster or more
slowly through the column depending on their properties
Different types of chromatography
column - separates by different properties of the protein
ion exchange - separates proteins by their net electric charge at a given pH
Cation exchange - the solid matrix has negatively charged groups
affinity - separates proteins by their binding affinity and specifications
size exclusion - separates protein by their size
How can high-performance liquid
chromatography (HPLC) improve the efficiency of chromatography methods?
it uses high pressure pumps that speed the movement of proteins down the column, and higher quality chromatographic materials. HPLC can limit diffusional spreading of protein bands and thus greatly improve resolution
What are the purification steps for an enzyme?
- Crude cellular extract
- Precipitation with ammonium sulfate
- Ion-exchange chromatography
- Size exclusion chromatography
- Affinity chromatography
What is the disadvantage of electrophoresis as a method to purify proteins and what it is used for alternatively?
it majorly affect the structure and function of the protein and there are other alternatives that do that; however, it is a great analytical tool to determine certain properties such as the protein’s isoelectric point and approximate molecular weight
Why is it not necessary to charge DNA with SDS for electrophoresis?
DNA is already negatively charged so it will not benefit from binding SDS which is used to add a net negative charge to the protein
In protein rich diet, which enzymes will be overly transcribed?
1. enzymes involved in glycolysis
2. enzymes involved in gluconeogenesis
3. enzymes involved in FA synthesis
4. enzymes involved in glycogen synthesis
- enzymes involved in gluconeogenesis
because the body doesn’t receive enough sugars from diet, so it uses AAs in proteins to generate glucose for energy.
Which AAs undergo phosphorylation as part of covalent control?
Serine, Threonine, Histidine, Tyrosine
How does waste from protein breakdown in muscles reach the liver?
Transamination of alanine.
The 2 types of nytrogenic waste come from 1. nucleic acid breakdown (in all cells) and 2. AA breakdown (in few tissues, mainly muscles). AA breakdown occurs in muscles to produce pyruvate to generate ATP and control cellular processes. most of the AA waste in the periphery reach the liver in the form of a different AAs- glutamine and alanine, where in the muscles alanine generation (transamination) is the preferred form, with the aid of the enzyme alanine aminotransferase.
