Three-Dimensional Structure of Carboxypeptidase T from Thermoactinomyces vulgaris in Complex with N-BOC-L-leucine

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V. I. Timofeev¹, S. A. Kuznetsov¹, V. Kh. Akparov², G. G. Chestukhina², and I. P. Kuranova^1*

¹Shubnikov Institute of Crystallography, Russian Academy of Sciences, Leninsky pr. 59, 119333 Moscow, Russia; fax: (499) 135-1011; E-mail: inna@ns.crys.ras.ru

²Scientific Center of Russian Federation Research Institute for Genetics and Selection of Industrial Microorganisms, 1-yi Dorozhnyi Proezd 1, 117545 Moscow, Russia; fax: (495) 315-0501; E-mail: valery@akparov.ru

^* To whom correspondence should be addressed.

Received October 15, 2012; Revision received November 9, 2012

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The 3D structure of recombinant bacterial carboxypeptidase T (CPT) in complex with N-BOC-L-leucine was determined at 1.38 Å resolution. Crystals for the X-ray study were grown in microgravity using the counter-diffusion technique. N-BOC-L-leucine and SO₄^2– ion bound in the enzyme active site were localized in the electron density map. Location of the leucine side chain in CPT–N-BOC-L-leucine complex allowed identification of the S1 subsite of the enzyme, and its structure was determined. Superposition of the structures of CPT–N-BOC-L-leucine complex and complexes of pancreatic carboxypeptidases A and B with substrate and inhibitors was carried out, and similarity of the S1 subsites in these three carboxypeptidases was revealed. It was found that SO₄^2– ion occupies the same position in the S1′ subsite as the C-terminal carboxy group of the substrate.

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KEY WORDS: carboxypeptidase T, S1 subsite, crystal structure, X-ray structural analysis

DOI: 10.1134/S0006297913030061

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