Question

3. A protein contains the following amino acids: O ALA 4 GLN 1 LEU 3 ARG 4 GLU 4 LYS 4 ASN 5 GLY O MET 1 ASP 1 HIS 2 PHE 4 ILE 4 PRO 8 SER 5 THR 1 TRP 2 TYR 2 VAL BGYS. a) What is its net charge at pH 1? b) What is its net charge at PH 13? c) Calculate the pl. 4. In what order would the amino acids GLU, LYS, and VAL be eluted from a cation-ion exchange column cs at a pH of 82 with CamScanner

Answer 1

Answer 3:

When a protein is formed, amino and carboxylic groups are involved in peptide bond formation. So only the presence of amino or carboxylic group in side chain of component amino acid units will determine the net charge.

In case of low pH (generally <3 ) NH2 gets protonated and converted to NH3+ and gains a positive charge

and at high pH (generally >10) C00H gets deprotonated to COO- and gains a negative charge

Analyzing the side groups from the image below we can infer:

NON-POLAR + CHARGE N ... H2N-C- --6 H 0 0 H.N-C-C CH HẠN*-c-cº H,N-C-C - H,Nº-c-c 0 . CH. ---- H,Nh-c-c CH, Học CH, CH, CH, CH SH 1-6-5--8- Glycine (Gly/G) Alanine (Ala/A) Valine (Val/V) Cysteine (Cys/C) Proline (Pro/P) C= NH+ NH2 H Lysine (Lys/K) Arginine (Arg/R) H 0 H H,Nº-c-c H.N-C-6 ,N-C-C CH, CH, o- H H,Nº-c-c.. CH, 0 0 H,Nº-c-c? H2N-C-C CH2 H.C-CHO CH CH NH CH, CH, CH, NH cine Leucine (Leu/L) Isoleucine (lle/I) Methionine (Met/M) Tryptophan (Trp/W) Phenylalanine (Phe/F) Histidine (His/H) POLAR - CHARGE H I-o 0 HN-C-C H,N*-C- C H.N- º- H,Nº- c o- CH, -o 0- CH, H,N*-¢-C I-0-0 H,N-C- C CH, o- 0 CH, CH, 0 H 2N-C-C CH 0 OH CH, No. OH NH, o Serine (Ser/S) Threonine (Thr/T) Tyrosine (Tyr/Y) Asparagine (Asn/N) Glutamine (Gin /Q) Aspartic Acid (Asp/D) Glutamic Acid (Glu/E)

3(a)

Given 4 GLN,3 ARG,4 LYS,4 ASN,1 HIS amino acids in protein

At pH 1(low pH)

so net charge = +1(4)+1(3)+1(4)+1(4)+1(1)= +16

As there is net positive charge of +1 per single amino acid molecule of GLN, ARG, LYS, ASN, HIS respectively

So, Net charge at protein at pH 1 is +16

3(b)

Given 4 GLU,1 ASP amino acids in protein

At pH 13(high pH)

so net charge = -1(4)-1(1) =-5

As there is net positive charge of -1 per single amino acid molecule of GLU, ASP respectively

So, Net charge at protein at pH 1 is -5

3(c) There are ionizable groups in this protein with pK values in increasing order as

3.86, 4.25, 6.04, 8.33, 10.07,10.79, 12.48 for ASP, GLU, HIS, CYS,TYR, LYS ARG respectively.

pI=(pK1+pK2) / 2

=7.18