Disulphide bond assignment in human tissue inhibitor of metalloproteinases (TIMP).
Disulphide bonds in human recombinant tissue inhibitor of metalloproteinases (TIMP) were assigned by resolving proteolytic digests of TIMP on reverse-phase h.p.l.c. and sequencing those peaks judged to contain disulphide bonds by virtue of a change in retention time on reduction. This procedure allowed the direct assignment of Cys-145-Cys-166 and the isolation of two other peptides containing two disulphide bonds each. Further peptide cleavage in conjunction with fast-atom-bombardment m.s. analysis permitted the assignments Cys-1-Cys-70, Cys-3-Cys-99, Cys-13-Cys-124 and Cys-127-Cys-174 from these peptides. The sixth bond Cys-132-Cys-137 was assigned by inference, as the native protein has no detectable free thiol groups.
Williamson
R A
RA
Biological Laboratory, University of Kent, Canterbury, U.K.
Marston
F A
FA
Angal
S
S
Koklitis
P
P
Panico
M
M
Morris
H R
HR
Carne
A F
AF
Smith
B J
BJ
Harris
T J
TJ
Freedman
R B
RB
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000032:analysis; D004220:Disulfides / Q000032:analysis*; D006023:Glycoproteins; D006801:Humans; D006868:Hydrolysis; D008666:Metalloendopeptidases; D051379:Mice; D003012:Microbial Collagenase / Q000037:antagonists & inhibitors; D008969:Molecular Sequence Data; D010434:Pepsin A; D011994:Recombinant Proteins / Q000235:genetics; D019714:Tissue Inhibitor of Metalloproteinases; D014357:Trypsin
D004220:Disulfides; D006023:Glycoproteins; D011994:Recombinant Proteins; D019714:Tissue Inhibitor of Metalloproteinases; D004247:DNA; D014357:Trypsin; D010434:Pepsin A; D008666:Metalloendopeptidases; D003012:Microbial Collagenase