1OS8

RECOMBINANT STREPTOMYCES GRISEUS TRYPSIN

Summary for 1OS8

• Entry DOI: 10.2210/pdb1os8/pdb
• Related: 1SGT
• Descriptor: trypsin, SULFATE ION, CALCIUM ION, ... (4 entities in total)
• Functional Keywords: trypsin, serine protease, hydrolase
• Biological source: Streptomyces griseus
• Total number of polymer chains: 1
• Total formula weight: 23258.93

Authors

Page, M.J.,Wong, S.L.,Hewitt, J.,Strynadka, N.C.,MacGillivray, R.T. (deposition date: 2003-03-18, release date: 2003-08-19, Last modification date: 2023-08-16)

Primary citation

Page, M.J.,Wong, S.L.,Hewitt, J.,Strynadka, N.C.,MacGillivray, R.T.
Engineering the Primary Substrate Specificity of Streptomyces griseus Trypsin.
Biochemistry, 42:9060-9066, 2003

PubMed Abstract: Streptomyces griseus trypsin (SGT) was chosen as a model scaffold for the development of serine proteases with enhanced substrate specificity. Recombinant SGT has been produced in a Bacillus subtilis expression system in a soluble active form and purified to homogeneity. The recombinant and native proteases have nearly identical enzymatic properties and structures. Four SGT mutants with alterations in the S1 substrate binding pocket (T190A, T190P, T190S, and T190V) were also expressed. The T190P mutant demonstrated the largest shift to a preference for Arg versus Lys in the P1 site. This was shown by a minor reduction in catalytic activity toward an Arg-containing substrate (k(cat) reduction of 25%). The crystal structures of the recombinant SGT and the T190P mutant in a complex with the inhibitor benzamidine were obtained at high resolution (approximately 1.9 A). The increase in P1 specificity, achieved with minimal effect on the catalytic efficiency, demonstrates that the T190P mutant is an ideal candidate for the design of additional substrate specificity engineered into the S2 to S4 binding pockets.

PubMed: 12885239
DOI: 10.1021/bi0344230

Experimental method

X-RAY DIFFRACTION (1.55 Å)