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Chaperone-assisted overexpression of an active D-carbamoylase from Agrobacterium tumefaciens AM 10.

DIR@IMTECH: CSIR-Institute of Microbial Technology

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Title Chaperone-assisted overexpression of an active D-carbamoylase from Agrobacterium tumefaciens AM 10.
 
Creator Sareen, D
Sharma, R
Vohra, R M
 
Subject QR Microbiology
 
Description The N-carbamoyl-D-amino acid amidohydrolase (D-carbamoylase) gene (dcb) from Agrobacterium tumefaciens AM 10 was cloned by polymerase chain reaction in plasmid pET28a and was overexpressed in Escherichia coli JM109 (DE3). However, almost 80% of the enzyme remained trapped in inclusion bodies. To facilitate the expression of the properly folded active enzyme, the chaperones GroEL/ES were coexpressed in plasmid pKY206. This resulted in a 43-fold increase in active enzyme production compared to the wild-type strain. The histidyl-tagged D-carbamoylase was purified by a single step nickel-affinity chromatography to a specific activity of 9.5 U/mg protein.
 
Publisher Elsevier Science
 
Date 2001-12
 
Type Article
PeerReviewed
 
Format application/pdf
 
Identifier http://crdd.osdd.net/open/282/1/vohra2001.pdf
Sareen, D and Sharma, R and Vohra, R M (2001) Chaperone-assisted overexpression of an active D-carbamoylase from Agrobacterium tumefaciens AM 10. Protein expression and purification, 23 (3). pp. 374-9. ISSN 1046-5928
 
Relation http://www.sciencedirect.com/science/article/pii/S1046592801915323
http://crdd.osdd.net/open/282/