2jev: Difference between revisions
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==Overview== | ==Overview== | ||
The N1-acetylation of spermidine and spermine by spermidine/spermine, acetyltransferase (SSAT) is a crucial step in the regulation of the, cellular polyamine levels in eukaryotic cells. Altered polyamine levels, are associated with a variety of cancers as well as other diseases, and, key enzymes in the polyamine pathway, including SSAT, are being explored, as potential therapeutic drug targets. We have expressed and purified, human SSAT in Escherichia coli and characterized its kinetic and chemical, mechanism. Initial velocity and inhibition studies support a random, sequential mechanism for the enzyme. The bisubstrate analogue, N1-spermine-acetyl-coenzyme A, exhibited linear, competitive inhibition, against both substrates with a true Ki of 6 nM. The pH-activity profile, was bell-shaped, depending on the ionization state of two groups, exhibiting apparent pKa values of 7.27 and 8.87. The three-dimensional, crystal structure of SSAT with bound bisubstrate inhibitor was determined, at 2.3 A resolution. The structure of the SSAT-spermine-acetyl-coenzyme A, complex suggested that Tyr140 acts as general acid and Glu92, through one, or more water molecules, acts as the general base during catalysis. On the, basis of kinetic properties, pH dependence, and structural information, we, propose an acid/base-assisted reaction catalyzed by SSAT, involving a, ternary complex. | The N1-acetylation of spermidine and spermine by spermidine/spermine, acetyltransferase (SSAT) is a crucial step in the regulation of the, cellular polyamine levels in eukaryotic cells. Altered polyamine levels, are associated with a variety of cancers as well as other diseases, and, key enzymes in the polyamine pathway, including SSAT, are being explored, as potential therapeutic drug targets. We have expressed and purified, human SSAT in Escherichia coli and characterized its kinetic and chemical, mechanism. Initial velocity and inhibition studies support a random, sequential mechanism for the enzyme. The bisubstrate analogue, N1-spermine-acetyl-coenzyme A, exhibited linear, competitive inhibition, against both substrates with a true Ki of 6 nM. The pH-activity profile, was bell-shaped, depending on the ionization state of two groups, exhibiting apparent pKa values of 7.27 and 8.87. The three-dimensional, crystal structure of SSAT with bound bisubstrate inhibitor was determined, at 2.3 A resolution. The structure of the SSAT-spermine-acetyl-coenzyme A, complex suggested that Tyr140 acts as general acid and Glu92, through one, or more water molecules, acts as the general base during catalysis. On the, basis of kinetic properties, pH dependence, and structural information, we, propose an acid/base-assisted reaction catalyzed by SSAT, involving a, ternary complex. | ||
==Disease== | |||
Known disease associated with this structure: Keratosis follicularis spinulosa decalvans OMIM:[[http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=313020 313020]] | |||
==About this Structure== | ==About this Structure== | ||
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[[Category: transferase]] | [[Category: transferase]] | ||
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Revision as of 23:48, 12 November 2007
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CRYSTAL STRUCTURE OF HUMAN SPERMINE,SPERMIDINE ACETYLTRANSFERASE IN COMPLEX WITH A BISUBSTRATE ANALOG (N1-ACETYLSPERMINE-S-COA).
OverviewOverview
The N1-acetylation of spermidine and spermine by spermidine/spermine, acetyltransferase (SSAT) is a crucial step in the regulation of the, cellular polyamine levels in eukaryotic cells. Altered polyamine levels, are associated with a variety of cancers as well as other diseases, and, key enzymes in the polyamine pathway, including SSAT, are being explored, as potential therapeutic drug targets. We have expressed and purified, human SSAT in Escherichia coli and characterized its kinetic and chemical, mechanism. Initial velocity and inhibition studies support a random, sequential mechanism for the enzyme. The bisubstrate analogue, N1-spermine-acetyl-coenzyme A, exhibited linear, competitive inhibition, against both substrates with a true Ki of 6 nM. The pH-activity profile, was bell-shaped, depending on the ionization state of two groups, exhibiting apparent pKa values of 7.27 and 8.87. The three-dimensional, crystal structure of SSAT with bound bisubstrate inhibitor was determined, at 2.3 A resolution. The structure of the SSAT-spermine-acetyl-coenzyme A, complex suggested that Tyr140 acts as general acid and Glu92, through one, or more water molecules, acts as the general base during catalysis. On the, basis of kinetic properties, pH dependence, and structural information, we, propose an acid/base-assisted reaction catalyzed by SSAT, involving a, ternary complex.
DiseaseDisease
Known disease associated with this structure: Keratosis follicularis spinulosa decalvans OMIM:[313020]
About this StructureAbout this Structure
2JEV is a Single protein structure of sequence from Homo sapiens with NHQ as ligand. Active as Diamine N-acetyltransferase, with EC number 2.3.1.57 Structure known Active Site: AC1. Full crystallographic information is available from OCA.
ReferenceReference
Mechanistic and structural analysis of human spermidine/spermine N1-acetyltransferase., Hegde SS, Chandler J, Vetting MW, Yu M, Blanchard JS, Biochemistry. 2007 Jun 19;46(24):7187-95. Epub 2007 May 22. PMID:17516632
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