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J Virol, April 1998, p. 3383-3386, Vol. 72, No. 4
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Efficient Expression and Rapid Purification of Human T-Cell Leukemia Virus Type 1 Protease

Y. Shirley Ding,¹ Sherry M. Owen,^1,2 Renu B. Lal,² and Richard A. Ikeda^1,*

School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta, Georgia 30332-0400,¹ and Retrovirus Diseases Branch, Division of Viral and Rickettsial Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 30333²

Received 24 September 1997/Accepted 12 December 1997

Human T-cell leukemia virus type 1 (HTLV-1) is an oncovirus that is clinically associated with adult T-cell leukemia. We report here the construction of a pET19-based expression clone containing HTLV-1 protease fused to a decahistidine-containing leader peptide. The recombinant protein is efficiently expressed in Escherichia coli, and the fusion protein can be easily purified by affinity chromatography. Active mature protease in yields in excess of 3 mg/liter of culture can then be obtained by a novel two-step refolding and autoprocessing procedure. The purified enzyme exhibited K_m and K_cat values of 0.3 mM and 0.143 sec¹ at pH 5.3 and was inhibited by pepstatin A.

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^* Corresponding author. Mailing address: School of Chemistry and Biochemistry, Georgia Institute of Technology, 706 State St., Atlanta, GA 30332-0400. Phone: (404) 894-4037. Fax: (404) 894-7452. E-mail: RICK.IKEDA{at}CHEMISTRY.GATECH.EDU.

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