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Kinetic and Structural Impact of Metal Ions and Genetic Variations on Human DNA Polymerase iota
- Kinetic and Structural Impact of Metal Ions and Genetic Variations on Human DNA Polymerase iota
- Choi, JY[Choi, Jeong-Yun]; Patra, A[Patra, Amritaj]; Yeom, M[Yeom, Mina]; Lee, YS[Lee, Young-Sam]; Zhang, QQ[Zhang, Qianqian]; Egli, M[Egli, Martin]; Guengerich, FP[Guengerich, F. Peter]
- DGIST Authors
- Lee, YS[Lee, Young-Sam]
- Issue Date
- Journal of Biological Chemistry, 291(40), 21063-+
- Article Type
- Amino Acids; Catalysis; Catalyst Activity; Catalytic Functions; Cation-Pi Interactions; Chains; Coordination Reactions; Distance Geometry; Genes; Human Dna Polymerase; Manganese; Metal Ions; Metals; Octahedral Coordination Geometry; Polymers; Structural Effect; Structural Impact; Translesion Synthesis
- DNA polymerase (pol) ι is a Y-family polymerase involved in translesion synthesis, exhibiting higher catalytic activity with Mn2+ than Mg2+. The human germline R96G variant impairs both Mn2+-dependent and Mg2+-dependent activities of pol ι, whereas the Δ1-25 variant selectively enhances its Mg2+-dependent activity. We analyzed pre-steady-state kinetic and structural effects of these two metal ions and genetic variations on pol ι using pol ι core (residues 1-445) proteins. The presence of Mn2+ (0.15 mM) instead of Mg2+ (2 mM) caused a 770-fold increase in efficiency (kpol/Kd,dCTP) of pol ι for dCTP insertion opposite G, mainly due to a 450-fold decrease in Kd,dCTP. The R96G and Δ1-25 variants displayed a 53-fold decrease and a 3-fold increase, respectively, in kpol/Kd,dCTP for dCTP insertion opposite G with Mg2+ when compared with wild type, substantially attenuated by substitution with Mn2+. Crystal structures of pol ι ternary complexes, including the primer terminus 3′-OH and a non-hydrolyzable dCTP analogue opposite G with the active-site Mg2+ or Mn2+, revealed that Mn2+ achieves more optimal octahedral coordination geometry than Mg2+, with lower values in average coordination distance geometry in the catalytic metal A-site. Crystal structures of R96G revealed the loss of three H-bonds of residues Gly-96 and Tyr-93 with an incoming dNTP, due to the lack of an arginine, as well as a destabilized Tyr-93 side chain secondary to the loss of a cation-π interaction between both side chains. These results provide a mechanistic basis for alteration in pol ι catalytic function with coordinating metals and genetic variation. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.
- American Society for Biochemistry and Molecular Biology Inc.
- Related Researcher
Lee, Young Sam
Lab of genome maintenance
DNA replication and repair; Restoration of cellular senescence; Structural and functional relationship of proteins
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