CMP kinase from Escherichia coli is structurally related to other nucleoside monophosphate kinases.
CMP kinase from Escherichia coli is a monomeric protein of 225 amino acid residues. The protein exhibits little overall sequence similarities with other known NMP kinases. However, residues involved in binding of substrates and/or in catalysis were found conserved, and sequence comparison suggested conservation of the global fold found in adenylate kinases or in several CMP/UMP kinases. The enzyme was purified to homogeneity, crystallized, and analyzed for its structural and catalytic properties. The crystals belong to the hexagonal space group P6(3), have unit cell parameters a = b = 82.3 A and c = 60.7 A, and diffract x-rays to a 1.9 A resolution. The bacterial enzyme exhibits a fluorescence emission spectrum with maximum at 328 nm upon excitation at 295 nm, which suggests that the single tryptophan residue (Trp30) is located in a hydrophobic environment. Substrate specificity studies showed that CMP kinase from E. coli is active with ATP, dATP, or GTP as donors and with CMP, dCMP, and arabinofuranosyl-CMP as acceptors. This is in contrast with CMP/UMP kinase from Dictyostelium discoideum, an enzyme active on CMP or UMP but much less active on the corresponding deoxynucleotides. Binding of CMP enhanced the affinity of E. coli CMP kinase for ATP or ADP, a particularity never described in this family of proteins that might explain inhibition of enzyme activity by excess of nucleoside monophosphate.
Bucurenci
N
N
Unité de Biochimie des Régulations Cellulaires, Institut Pasteur, Paris, France.
Sakamoto
H
H
Briozzo
P
P
Palibroda
N
N
Serina
L
L
Sarfati
R S
RS
Labesse
G
G
Briand
G
G
Danchin
A
A
Bărzu
O
O
Gilles
A M
AM
D000595:Amino Acid Sequence; D002384:Catalysis; D003460:Crystallization; D004926:Escherichia coli / Q000201:enzymology*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009703:Nucleoside-Phosphate Kinase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013050:Spectrometry, Fluorescence; D014961:X-Ray Diffraction
C019979:cytidylate kinase; D009703:Nucleoside-Phosphate Kinase