1os8: Difference between revisions
New page: left|200px<br /><applet load="1os8" size="450" color="white" frame="true" align="right" spinBox="true" caption="1os8, resolution 1.55Å" /> '''RECOMBINANT STREPTOM... |
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[[Image:1os8.jpg|left|200px]]<br /><applet load="1os8" size=" | [[Image:1os8.jpg|left|200px]]<br /><applet load="1os8" size="350" color="white" frame="true" align="right" spinBox="true" | ||
caption="1os8, resolution 1.55Å" /> | caption="1os8, resolution 1.55Å" /> | ||
'''RECOMBINANT STREPTOMYCES GRISEUS TRYPSIN'''<br /> | '''RECOMBINANT STREPTOMYCES GRISEUS TRYPSIN'''<br /> | ||
==Overview== | ==Overview== | ||
Streptomyces griseus trypsin (SGT) was chosen as a model scaffold for the | 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. | ||
==About this Structure== | ==About this Structure== | ||
1OS8 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Streptomyces_chryseus Streptomyces chryseus] with SO4 and CA as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Trypsin Trypsin], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.4.21.4 3.4.21.4] Full crystallographic information is available from [http:// | 1OS8 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Streptomyces_chryseus Streptomyces chryseus] with <scene name='pdbligand=SO4:'>SO4</scene> and <scene name='pdbligand=CA:'>CA</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Trypsin Trypsin], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.4.21.4 3.4.21.4] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1OS8 OCA]. | ||
==Reference== | ==Reference== | ||
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[[Category: Trypsin]] | [[Category: Trypsin]] | ||
[[Category: Hewitt, J.]] | [[Category: Hewitt, J.]] | ||
[[Category: MacGillivray, R | [[Category: MacGillivray, R T.]] | ||
[[Category: Page, M | [[Category: Page, M J.]] | ||
[[Category: Strynadka, N | [[Category: Strynadka, N C.]] | ||
[[Category: Wong, S | [[Category: Wong, S L.]] | ||
[[Category: CA]] | [[Category: CA]] | ||
[[Category: SO4]] | [[Category: SO4]] | ||
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[[Category: trypsin]] | [[Category: trypsin]] | ||
''Page seeded by [http:// | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 14:21:01 2008'' | ||
Revision as of 15:21, 21 February 2008
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RECOMBINANT STREPTOMYCES GRISEUS TRYPSIN
OverviewOverview
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.
About this StructureAbout this Structure
1OS8 is a Single protein structure of sequence from Streptomyces chryseus with and as ligands. Active as Trypsin, with EC number 3.4.21.4 Full crystallographic information is available from OCA.
ReferenceReference
Engineering the primary substrate specificity of Streptomyces griseus trypsin., Page MJ, Wong SL, Hewitt J, Strynadka NC, MacGillivray RT, Biochemistry. 2003 Aug 5;42(30):9060-6. PMID:12885239
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