2on6: Difference between revisions

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==Overview==

The X-ray crystal structures of human purine nucleoside phosphorylase, (PNP) with bound inosine or transition-state analogues show His257 within, hydrogen bonding distance of the 5'-hydroxyl. The mutants His257Phe, His257Gly, and His257Asp exhibited greatly decreased affinity for, Immucillin-H (ImmH), binding this mimic of an early transition state as, much as 370-fold (Km/Ki) less tightly than native PNP. In contrast, these, mutants bound DADMe-ImmH, a mimic of a late transition state, nearly as, well as the native enzyme. These results indicate that His257 serves an, important role in the early stages of transition-state formation. Whereas, mutation of His257 resulted in little variation in the PNP.DADMe-ImmH.SO4, structures, His257Phe.ImmH.PO4 showed distortion at the 5'-hydroxyl, indicating the importance of H-bonding in positioning this group during, progression to the transition state. Binding isotope effect (BIE) and, kinetic isotope effect (KIE) studies of the remote 5'-3H for the, arsenolysis of inosine with native PNP revealed a BIE of 1.5% and an, unexpectedly large intrinsic KIE of 4.6%. This result is interpreted as a, moderate electronic distortion toward the transition state in the, Michaelis complex with continued development of a similar distortion at, the transition state. The mutants His257Phe, His257Gly, and His257Asp, altered the 5'-3H intrinsic KIE to -3, -14, and 7%, respectively, while, the BIEs contributed 2, 2, and -2%, respectively. These surprising results, establish that forces in the Michaelis complex, reported by the BIEs, can, be reversed or enhanced at the transition state.

The X-ray crystal structures of human purine nucleoside phosphorylase (PNP) with bound inosine or transition-state analogues show His257 within hydrogen bonding distance of the 5'-hydroxyl. The mutants His257Phe, His257Gly, and His257Asp exhibited greatly decreased affinity for Immucillin-H (ImmH), binding this mimic of an early transition state as much as 370-fold (Km/Ki) less tightly than native PNP. In contrast, these mutants bound DADMe-ImmH, a mimic of a late transition state, nearly as well as the native enzyme. These results indicate that His257 serves an important role in the early stages of transition-state formation. Whereas mutation of His257 resulted in little variation in the PNP x DADMe-ImmH x SO4 structures, His257Phe x ImmH x PO4 showed distortion at the 5'-hydroxyl, indicating the importance of H-bonding in positioning this group during progression to the transition state. Binding isotope effect (BIE) and kinetic isotope effect (KIE) studies of the remote 5'-(3)H for the arsenolysis of inosine with native PNP revealed a BIE of 1.5% and an unexpectedly large intrinsic KIE of 4.6%. This result is interpreted as a moderate electronic distortion toward the transition state in the Michaelis complex with continued development of a similar distortion at the transition state. The mutants His257Phe, His257Gly, and His257Asp altered the 5'-(3)H intrinsic KIE to -3, -14, and 7%, respectively, while the BIEs contributed 2, 2, and -2%, respectively. These surprising results establish that forces in the Michaelis complex, reported by the BIEs, can be reversed or enhanced at the transition state.

==Disease==

Known diseases associated with this structure: Neutral lipid storage disease with myopathy OMIM:[[http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=609059 609059]], Nucleoside phosphorylase deficiency, immunodeficiency due to OMIM:[[http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=164050 164050]]

==About this Structure==

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==Reference==

Neighboring ~~Group Participation~~ in the ~~Transition State~~ of ~~Human Purine Nucleoside Phosphorylase~~., Murkin AS, Birck MR, Rinaldo-Matthis A, Shi W, Taylor EA ~~Steven C~~ Almo, Schramm VL, Biochemistry. 2007 May 1;46(17):5038-~~5049~~. Epub 2007 Apr 4. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=17407325 17407325]

Neighboring group participation in the transition state of human purine nucleoside phosphorylase., Murkin AS, Birck MR, Rinaldo-Matthis A, Shi W, Taylor EA, Almo SC, Schramm VL, Biochemistry. 2007 May 1;46(17):5038-49. Epub 2007 Apr 4. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=17407325 17407325]

[[Category: Homo sapiens]]

[[Category: Purine-nucleoside phosphorylase]]

[[Category: Single protein]]

[[Category: Almo, S.C.]]

[[Category: Almo, S C.]]

[[Category: Murkin, A.S.]]

[[Category: Murkin, A S.]]

[[Category: Rinaldo-Matthis, A.]]

[[Category: Schramm, V.L.]]

[[Category: Schramm, V L.]]

[[Category: IMH]]

[[Category: purine nucleoside phosphorylase]]

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''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 18:20:18 2008''

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@@ Line 4: / Line 4: @@
 ==Overview==
-The X-ray crystal structures of human purine nucleoside phosphorylase, (PNP) with bound inosine or transition-state analogues show His257 within, hydrogen bonding distance of the 5'-hydroxyl. The mutants His257Phe, His257Gly, and His257Asp exhibited greatly decreased affinity for, Immucillin-H (ImmH), binding this mimic of an early transition state as, much as 370-fold (Km/Ki) less tightly than native PNP. In contrast, these, mutants bound DADMe-ImmH, a mimic of a late transition state, nearly as, well as the native enzyme. These results indicate that His257 serves an, important role in the early stages of transition-state formation. Whereas, mutation of His257 resulted in little variation in the PNP.DADMe-ImmH.SO4, structures, His257Phe.ImmH.PO4 showed distortion at the 5'-hydroxyl, indicating the importance of H-bonding in positioning this group during, progression to the transition state. Binding isotope effect (BIE) and, kinetic isotope effect (KIE) studies of the remote 5'-3H for the, arsenolysis of inosine with native PNP revealed a BIE of 1.5% and an, unexpectedly large intrinsic KIE of 4.6%. This result is interpreted as a, moderate electronic distortion toward the transition state in the, Michaelis complex with continued development of a similar distortion at, the transition state. The mutants His257Phe, His257Gly, and His257Asp, altered the 5'-3H intrinsic KIE to -3, -14, and 7%, respectively, while, the BIEs contributed 2, 2, and -2%, respectively. These surprising results, establish that forces in the Michaelis complex, reported by the BIEs, can, be reversed or enhanced at the transition state.
+The X-ray crystal structures of human purine nucleoside phosphorylase (PNP) with bound inosine or transition-state analogues show His257 within hydrogen bonding distance of the 5'-hydroxyl. The mutants His257Phe, His257Gly, and His257Asp exhibited greatly decreased affinity for Immucillin-H (ImmH), binding this mimic of an early transition state as much as 370-fold (Km/Ki) less tightly than native PNP. In contrast, these mutants bound DADMe-ImmH, a mimic of a late transition state, nearly as well as the native enzyme. These results indicate that His257 serves an important role in the early stages of transition-state formation. Whereas mutation of His257 resulted in little variation in the PNP x DADMe-ImmH x SO4 structures, His257Phe x ImmH x PO4 showed distortion at the 5'-hydroxyl, indicating the importance of H-bonding in positioning this group during progression to the transition state. Binding isotope effect (BIE) and kinetic isotope effect (KIE) studies of the remote 5'-(3)H for the arsenolysis of inosine with native PNP revealed a BIE of 1.5% and an unexpectedly large intrinsic KIE of 4.6%. This result is interpreted as a moderate electronic distortion toward the transition state in the Michaelis complex with continued development of a similar distortion at the transition state. The mutants His257Phe, His257Gly, and His257Asp altered the 5'-(3)H intrinsic KIE to -3, -14, and 7%, respectively, while the BIEs contributed 2, 2, and -2%, respectively. These surprising results establish that forces in the Michaelis complex, reported by the BIEs, can be reversed or enhanced at the transition state.
+==Disease==
+Known diseases associated with this structure: Neutral lipid storage disease with myopathy OMIM:[[http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=609059 609059]], Nucleoside phosphorylase deficiency, immunodeficiency due to OMIM:[[http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=164050 164050]]
 ==About this Structure==
@@ Line 10: / Line 13: @@
 ==Reference==
-Neighboring Group Participation in the Transition State of Human Purine Nucleoside Phosphorylase., Murkin AS, Birck MR, Rinaldo-Matthis A, Shi W, Taylor EA Steven C Almo, Schramm VL, Biochemistry. 2007 May 1;46(17):5038-5049. Epub 2007 Apr 4. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=17407325 17407325]
+Neighboring group participation in the transition state of human purine nucleoside phosphorylase., Murkin AS, Birck MR, Rinaldo-Matthis A, Shi W, Taylor EA, Almo SC, Schramm VL, Biochemistry. 2007 May 1;46(17):5038-49. Epub 2007 Apr 4. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=17407325 17407325]
 [[Category: Homo sapiens]]
 [[Category: Purine-nucleoside phosphorylase]]
 [[Category: Single protein]]
-[[Category: Almo, S.C.]]
+[[Category: Almo, S C.]]
-[[Category: Murkin, A.S.]]
+[[Category: Murkin, A S.]]
 [[Category: Rinaldo-Matthis, A.]]
-[[Category: Schramm, V.L.]]
+[[Category: Schramm, V L.]]
 [[Category: IMH]]
 [[Category: purine nucleoside phosphorylase]]
-''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Wed Jan 23 15:12:32 2008''
+''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 18:20:18 2008''