Overexpression, purification and crystallographic analysis of a unique adenosine kinase from Mycobacterium tuberculosis.
Adenosine kinase from Mycobacterium tuberculosis is the only prokaryotic adenosine kinase that has been isolated and characterized. The enzyme catalyzes the phosphorylation of adenosine to adenosine monophosphate and is involved in the activation of 2-methyladenosine, a compound that has demonstrated selective activity against M. tuberculosis. The mechanism of action of 2-methyladenosine is likely to be different from those of current tuberculosis treatments and this compound (or other adenosine analogs) may prove to be a novel therapeutic intervention for this disease. The M. tuberculosis adenosine kinase was overexpressed in Escherichia coli and the enzyme was purified with activity comparable to that reported previously. The protein was crystallized in the presence of adenosine using the vapour-diffusion method. The crystals diffracted X-rays to high resolution and a complete data set was collected to 2.2 A using synchrotron radiation. The crystal belonged to space group P3(1)21, with unit-cell parameters a = 70.2, c = 111.6 A, and contained a single protein molecule in the asymmetric unit. An initial structural model of the protein was obtained by the molecular-replacement method, which revealed a dimeric structure. The monomers of the dimer were related by twofold crystallographic symmetry. An understanding of how the M. tuberculosis adenosine kinase differs from the human homolog should aid in the design of more potent and selective antimycobacterial agents that are selectively activated by this enzyme.
Wang
Yimin
Y
Southern Research Institute, Birmingham, Alabama 35205, USA.
Long
Mary C
MC
Ranganathan
Senthil
S
Escuyer
Vincent
V
Parker
William B
WB
Li
Rongbao
R
D000248:Adenosine Kinase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D003001:Cloning, Molecular; D003460:Crystallization / Q000379:methods; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D014835:Volatilization; D014961:X-Ray Diffraction
D001426:Bacterial Proteins; D000248:Adenosine Kinase