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Expression and purification of a cold-adapted group III trypsin in Escherichia coli.

Abstract：

:The recently classified group III trypsins include members like Atlantic cod (Gadus morhua) trypsin Y as well as seven analogues from other cold-adapted fish species. The eight group III trypsins have been characterized from their cDNAs and deduced amino acid sequences but none of the enzymes have been isolated from their native sources. This study describes the successful expression and purification of a recombinant HP-thioredoxin-trypsin Y fusion protein in the His-Patch ThioFusion Escherichia coli expression system and its purification by chromatographic methods. The recombinant form of trypsin Y was previously expressed in Pichia pastoris making it the first biochemically characterized group III trypsin. It has dual substrate specificity towards trypsin and chymotrypsin substrates and demonstrates an increasing activity at temperatures between 2 and 21 degrees C with a complete inactivation at 30 degrees C. The aim of the study was to facilitate further studies of recombinant trypsin Y by finding an expression system yielding higher amounts of the enzyme than possible in our hands in the P. pastoris system. Also, commercial production of trypsin Y will require an efficient and inexpensive expression system like the His-Patch ThioFusion E. coli expression system described here as the enzyme is produced in very low amounts in the Atlantic cod.

journal_name

Protein Expr Purif

journal_title

Protein expression and purification

authors

Pálsdóttir HM,Gudmundsdóttir A

doi

10.1016/j.pep.2006.06.008

subject

Has Abstract

pub_date

2007-02-01 00:00:00

pages

243-52

issue

2

eissn

1046-5928

issn

1096-0279

pii

S1046-5928(06)00181-1

journal_volume

51

pub_type

杂志文章

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