Expression of human cathepsin K in Pichia pastoris and preliminary crystallographic studies of an inhibitor complex

✍ Scribed by Christopher J. Linnevers; Mary E. Mcgrath; Andy Armstrong; Firoz R. Mistry; Michael G. Barnes; Jeffrey L. Klaus; James T. Palmer; Bradley A. Katz; Dieter Brömme

• Book ID: 105356462
• Publisher: Cold Spring Harbor Laboratory Press
• Year: 2008
• Tongue: English
• Weight: 322 KB
• Volume: 6
• Category: Article
• ISSN: 0961-8368
• DOI: 10.1002/pro.5560060421

✦ Synopsis

Abstract

Cathepsin K is a cysteine protease of the papain family, which is predominantly expressed in osteoclasts, and is regarded as a key protease in bone remodeling. To facilitate structural studies of the protein, the wild‐type sequence of the protease has been mutated so as to replace a potential N‐glycosylation site. We have expressed the mutant human cathepsin K to 190 mg/5 L using the Pichia pastoris expression system. Cathepsin K was inactivated with the mechanism‐based inhibitor, APC3328, and crystallized from magnesium formate. A 2.2 Å X‐ray data set has been collected on crystals belonging to space group P2~1~2~1~2~1~, with a = 41.66 Å, b = 51.41 Å, and c = 107.72 Å. There is most likely one molecule per asymmetric unit.