1997
DOI: 10.1021/bi971609o
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Determinants of Ribose Specificity in RNA Polymerization:  Effects of Mn2+ and Deoxynucleoside Monophosphate Incorporation into Transcripts

Abstract: The catalytic specificity of T7 RNA polymerase (RNAP) for ribonucleoside triphosphates vs deoxynucleoside triphosphates {(kcat/Km)rNTP/(kcat/Km)dNTP} during transcript elongation is approximately 80. Mutation of tyrosine 639 to phenylalanine reduces specificity by a factor of approximately 20 and largely eliminates the Km difference between rNTPs and dNTPs. The remaining specificity factor of approximately 4 is kcat-mediated and is nearly eliminated if Mn2+ is substituted for Mg2+ in the reaction. Mn2+ substit… Show more

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Cited by 53 publications
(55 citation statements)
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“…Manganese is known to decrease the specificity for nucleotide incorporation by polymerases (10,34). Consistent with the effects on DNA-dependent RNA polymerases, we found Mn 2ϩ to slightly increase initiation from a 3Ј uridylate and expand the use of several GTP analogs for initiation (30).…”
Section: Vol 76 2002 Metals and De Novo Initiation Of Viral Rna Synsupporting
confidence: 82%
See 1 more Smart Citation
“…Manganese is known to decrease the specificity for nucleotide incorporation by polymerases (10,34). Consistent with the effects on DNA-dependent RNA polymerases, we found Mn 2ϩ to slightly increase initiation from a 3Ј uridylate and expand the use of several GTP analogs for initiation (30).…”
Section: Vol 76 2002 Metals and De Novo Initiation Of Viral Rna Synsupporting
confidence: 82%
“…Divalent metals are known to modulate substrate selectivity of DNA-dependent RNA polymerases (10,34). For the HCV RdRp, Mn 2ϩ improved the K m value for the initiation GTP from 103 to 3 M. Mn 2ϩ also decreased primer extension since its addition to reactions containing Mg 2ϩ not only increased de novo initiation but also reduced primer extension activity.…”
Section: Discussionmentioning
confidence: 99%
“…In addition, the template specificity of an RNA polymerase and the product length synthesized by these enzymes are their distinctive "fingerprint." The T7 RNA polymerase synthesizes poly(rG) five times more efficiently than poly(rA) on homopolymeric templates (37). Mcm10p synthesized poly(rA) 10 times more efficiently than poly(rG) (Fig.…”
Section: Discussionmentioning
confidence: 99%
“…An effect on metal binding would also account for the decreased catalytic activity of the enzyme and the decreased replication capacity of viruses harboring these mutations (7). Mn 2ϩ is compatible with a wider range of reactive group geometries than Mg 2ϩ (35) and thus Mn 2ϩ -catalyzed IN reactions and inhibition may be less sensitive to changes in reactive group geometries.…”
Section: Discussionmentioning
confidence: 99%