Open AccessThe expression and purification of human rhinovirus protease 3C
Author(s) -
KNOTT Jacqueline A.,
ORR David C.,
MONTGOMERY Douglas S.,
SULLIVAN Carole A.,
WESTON Anthony
Publication year - 1989
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1989.tb14862.x
Subject(s) - rhinovirus , protease , virology , expression (computer science) , biology , virus , enzyme , computer science , biochemistry , programming language
Human rhinovirus type 14 protease 3C was expressed as a soluble and active protein in Escherichia coli . The protease was purified by a cationic‐exchange step followed by gel filtration on a TSK 3000 column. The final yield of purified protease was in the range 0.5–1.0 mg/l culture grown to A 550 = 1.0. Sequence analysis revealed that greater than 90% of the N‐terminal residues were methionine. The enzyme activity of the purified protease was measured by cleavage of a synthetic peptide representing a predicted Gln/Gly viral polyprotein cleavage site. A mutant protease (Cys146→Ser) was produced and purified in the same way. The yield of mutant protease 3C was approximately 150 μg/l from a culture grown to A 550 = 1.0. This mutant protease 3C did not cleave the synthetic peptide substrate.