lecture 12 - disulphide bonds

Question 1

what is a disulphide bond?

Answer 1

• oxidized cys

- links together separate parts of a polypeptide

Question 2

how do we prepare a protein with a disulphide bond for sequencing?

Answer 2

• reduce cys-cys
• block SH groups (e.g. using carbomethylate)
• prevents reoxidation

Question 3

what is a con of how we sequence disulphide bonds

Answer 3

-destroys the disulphide connectivity

Question 4

how do we sequence protein with a disulphide bond?

2

Answer 4

(1) sequence full reduced protein OR
(2) keep the protein oxidized
- then cut the protein with enzyme
- isolate fragments
- sequence fragments OR test amino acid composition OR use mass spectrometry to get mass of fragments