Cloning, expression and mutational studies of a trypsin inhibitor that retains activity even after cyanogen bromide digestion.
Protein Expr Purif
; 96: 26-31, 2014 Apr.
Article
in En
| MEDLINE
| ID: mdl-24492011
ABSTRACT
A winged bean trypsin inhibitor (WbTI-2) of molecular mass â¼20kDa, has been cloned and expressed in Escherichiacoli with full activity like the one from seed protein. It completely inhibits trypsin at an enzymeinhibitor molar ratio of 12. PCR with cDNA and genomic DNA using same primers produced about 550 base pair product, which indicated it to be an intronless gene. Through site-directed mutagenesis, the Arg64 has been confirmed as the P1 residue. For the presence of five methionine residues in WbTI-2, cyanogen bromide (CNBr) digestion was carried out. Out of three fragments the one (about 65% of original size) containing the reactive site loop retained 50% activity.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Plant Proteins
/
Escherichia coli
Language:
En
Journal:
Protein Expr Purif
Journal subject:
BIOLOGIA MOLECULAR
Year:
2014
Type:
Article
Affiliation country:
India