2r7c: Difference between revisions

Revision as of 19:45, 21 February 2008

Crystallographic and biochemical analysis of rotavirus NSP2 with nucleotides reveals an NDP kinase like activity

OverviewOverview

Rotavirus, the major pathogen of infantile gastroenteritis, carries a nonstructural protein, NSP2, essential for viroplasm formation and genome replication/packaging. In addition to RNA-binding and helix-destabilizing properties, NSP2 exhibits nucleoside triphosphatase activity. A conserved histidine (H225) functions as the catalytic residue for this enzymatic activity, and mutation of this residue abrogates genomic double-stranded RNA synthesis without affecting viroplasm formation. To understand the structural basis of the phosphatase activity of NSP2, we performed crystallographic analyses of native NSP2 and a functionally defective H225A mutant in the presence of nucleotides. These studies showed that nucleotides bind inside a cleft between the two domains of NSP2 in a region that exhibits structural similarity to ubiquitous cellular HIT (histidine triad) proteins. Only minor conformational alterations were observed in the cleft upon nucleotide binding and hydrolysis. This hydrolysis involved the formation of a stable phosphohistidine intermediate. These observations, reminiscent of cellular nucleoside diphosphate (NDP) kinases, prompted us to investigate whether NSP2 exhibits phosphoryl-transfer activity. Bioluminometric assay showed that NSP2 exhibits an NDP kinase-like activity that transfers the bound phosphate to NDPs. However, NSP2 is distinct from the highly conserved cellular NDP kinases in both its structure and catalytic mechanism, thus making NSP2 a potential target for antiviral drug design. With structural similarities to HIT proteins, which are not known to exhibit NDP kinase activity, NSP2 represents a unique example among structure-activity relationships. The newly observed phosphoryl-transfer activity of NSP2 may be utilized for homeostasis of nucleotide pools in viroplasms during genome replication.

About this StructureAbout this Structure

2R7C is a Single protein structure of sequence from Simian rotavirus a/sa11 with as ligand. Full crystallographic information is available from OCA.

ReferenceReference

Crystallographic and biochemical analysis of rotavirus NSP2 with nucleotides reveals a nucleoside diphosphate kinase-like activity., Kumar M, Jayaram H, Vasquez-Del Carpio R, Jiang X, Taraporewala ZF, Jacobson RH, Patton JT, Prasad BV, J Virol. 2007 Nov;81(22):12272-84. Epub 2007 Sep 5. PMID:17804496

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 ==Overview==
-Rotavirus, the major pathogen of infantile gastroenteritis, encodes a, nonstructural protein NSP2 essential for viroplasm formation and genome, replication/packaging. In addition to RNA binding and helix-destabilizing, properties, NSP2 exhibits nucleoside triphosphatase activity. A conserved, histidine (H225) functions as the catalytic residue for this enzymatic, activity, and mutation of this residue abrogates genomic dsRNA synthesis, without affecting viroplasm formation. To understand the structural basis, of the phosphatase activity of NSP2 we performed crystallographic analyses, of native NSP2 and a functionally defective H225A mutant in the presence, of nucleotides. These studies showed that nucleotides bind inside a cleft, between the two domains of NSP2 in a region that exhibits structural, similarity to ubiquitous cellular HIT (Histidine Triad) proteins. Only, minor conformational alterations were observed in the cleft upon, nucleotide binding and hydrolysis. This hydrolysis involved the formation, of a stable phosphohistidine intermediate. These observations, reminiscent, of cellular nucleoside diphosphate (NDP) kinases, prompted us to, investigate whether NSP2 exhibits phosphoryl-transfer activity., Bioluminometric assay showed that NSP2 exhibits an NDP kinase-like, activity that transfers the bound phosphate to NDPs. However, NSP2 is, distinct from the highly conserved cellular NDP kinases both in its, structure and catalytic mechanism, thus making NSP2 a potential target for, antiviral drug design. With structural similarities to HIT proteins, which, are not known to exhibit NDP kinase activity, NSP2 represents a unique, example in structure-activity relationship. The newly-observed, phosphoryl-transfer activity of NSP2 may be utilized for homeostasis of, nucleotide pools in viroplasms during genome replication.
+Rotavirus, the major pathogen of infantile gastroenteritis, carries a nonstructural protein, NSP2, essential for viroplasm formation and genome replication/packaging. In addition to RNA-binding and helix-destabilizing properties, NSP2 exhibits nucleoside triphosphatase activity. A conserved histidine (H225) functions as the catalytic residue for this enzymatic activity, and mutation of this residue abrogates genomic double-stranded RNA synthesis without affecting viroplasm formation. To understand the structural basis of the phosphatase activity of NSP2, we performed crystallographic analyses of native NSP2 and a functionally defective H225A mutant in the presence of nucleotides. These studies showed that nucleotides bind inside a cleft between the two domains of NSP2 in a region that exhibits structural similarity to ubiquitous cellular HIT (histidine triad) proteins. Only minor conformational alterations were observed in the cleft upon nucleotide binding and hydrolysis. This hydrolysis involved the formation of a stable phosphohistidine intermediate. These observations, reminiscent of cellular nucleoside diphosphate (NDP) kinases, prompted us to investigate whether NSP2 exhibits phosphoryl-transfer activity. Bioluminometric assay showed that NSP2 exhibits an NDP kinase-like activity that transfers the bound phosphate to NDPs. However, NSP2 is distinct from the highly conserved cellular NDP kinases in both its structure and catalytic mechanism, thus making NSP2 a potential target for antiviral drug design. With structural similarities to HIT proteins, which are not known to exhibit NDP kinase activity, NSP2 represents a unique example among structure-activity relationships. The newly observed phosphoryl-transfer activity of NSP2 may be utilized for homeostasis of nucleotide pools in viroplasms during genome replication.
 ==About this Structure==
@@ Line 10: / Line 10: @@
 ==Reference==
-Crystallographic and biochemical analysis of rotavirus NSP2 with nucleotides reveals an NDP kinase like activity., Kumar M, Jayaram H, Carpio RV, Jiang X, Taraporwala ZF, Jacobson RH, Patton JT, Prasad BV, J Virol. 2007 Sep 5;. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=17804496 17804496]
+Crystallographic and biochemical analysis of rotavirus NSP2 with nucleotides reveals a nucleoside diphosphate kinase-like activity., Kumar M, Jayaram H, Vasquez-Del Carpio R, Jiang X, Taraporewala ZF, Jacobson RH, Patton JT, Prasad BV, J Virol. 2007 Nov;81(22):12272-84. Epub 2007 Sep 5. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=17804496 17804496]
 [[Category: Simian rotavirus a/sa11]]
 [[Category: Single protein]]
 [[Category: Kumar, M.]]
-[[Category: Prasad, B.V.V.]]
+[[Category: Prasad, B V.V.]]
 [[Category: PO4]]
 [[Category: ndp kinase]]
@@ Line 23: / Line 23: @@
 [[Category: rotavirus]]
-''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Wed Jan 23 12:12:58 2008''
+''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 18:45:10 2008''