Structural and functional features of an NDP kinase from the hyperthermophile crenarchaeon Pyrobaculum aerophilum.
Nucleoside diphosphate (NDP) kinases are ubiquitous enzymes that transfer gamma-phosphates from nucleoside triphosphates to nucleoside diphosphates via a ping-pong mechanism. The important role of this large family of enzymes in controlling cellular functions and developmental processes along with their crystallizability has made them good candidates for structural studies. We recently determined the structure of an evolved version of an NDP kinase from Pyrobaculum aerophilum, an extreme thermophile. This NDP kinase has similarity to the 42 other NDP kinases deposited in the Protein Data Bank (PDB) but differs significantly in sequence, structure, and biophysical properties. The P. aerophilum NDP kinase sequence contains two unique segments not present in other NDP kinases, comprising residues 66-100 and 156-165. We show that deletion mutants of the P. aerophilum NDP kinase lacking either or both of these inserts have an altered substrate specificity, allowing dGTP as the phosphate donor. A structural analysis of the evolved NDP kinase in conjunction with mutagenesis experiments suggests that the substrate specificity of the P. aerophilum NDP kinase is related to the presence of these two inserts.
Pédelacq
Jean-Denis
JD
Bioscience Division, MS-M888, Los Alamos National Laboratory, Los Alamos, NM 87545, USA. jpdlcq@lanl.gov
Waldo
Geoffrey S
GS
Cabantous
Stéphanie
S
Liong
Elaine C
EC
Terwilliger
Thomas C
TC
D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003848:Deoxyguanine Nucleotides / Q000378:metabolism; D019143:Evolution, Molecular; D009701:Nucleoside-Diphosphate Kinase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010710:Phosphates / Q000378:metabolism; D017434:Protein Structure, Tertiary; D041543:Pyrobaculum / Q000201:enzymology* / Q000235:genetics; D017384:Sequence Deletion / Q000235:genetics; D040681:Structural Homology, Protein; D013379:Substrate Specificity / Q000235:genetics
D019843:Archaeal Proteins; D003848:Deoxyguanine Nucleotides; D010710:Phosphates; C029603:deoxyguanosine triphosphate; D009701:Nucleoside-Diphosphate Kinase