Rifampin phosphotransferase is an unusual antibiotic resistance kinase.
Rifampin (RIF) phosphotransferase (RPH) confers antibiotic resistance by conversion of RIF and ATP, to inactive phospho-RIF, AMP and Pi. Here we present the crystal structure of RPH from Listeria monocytogenes (RPH-Lm), which reveals that the enzyme is comprised of three domains: two substrate-binding domains (ATP-grasp and RIF-binding domains); and a smaller phosphate-carrying His swivel domain. Using solution small-angle X-ray scattering and mutagenesis, we reveal a mechanism where the swivel domain transits between the spatially distinct substrate-binding sites during catalysis. RPHs are previously uncharacterized dikinases that are widespread in environmental and pathogenic bacteria. These enzymes are members of a large unexplored group of bacterial enzymes with substrate affinities that have yet to be fully explored. Such an enzymatically complex mechanism of antibiotic resistance augments the spectrum of strategies used by bacteria to evade antimicrobial compounds.
Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, Ontario, Canada M5G 1L6.
Peter J
PJ
Stogios
M.G. DeGroote Institute for Infectious Disease Research, Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main St W, Hamilton, Ontario, Canada L8S 4K1.
Georgina
G
Cox
M.G. DeGroote Institute for Infectious Disease Research, Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main St W, Hamilton, Ontario, Canada L8S 4K1.
Peter
P
Spanogiannopoulos
Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main St W, Hamilton, Ontario, Canada L8S 4K1.
Monica C
MC
Pillon
M.G. DeGroote Institute for Infectious Disease Research, Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main St W, Hamilton, Ontario, Canada L8S 4K1.
Nicholas
N
Waglechner
M.G. DeGroote Institute for Infectious Disease Research, Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main St W, Hamilton, Ontario, Canada L8S 4K1.
Tatiana
T
Skarina
M.G. DeGroote Institute for Infectious Disease Research, Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main St W, Hamilton, Ontario, Canada L8S 4K1.
Kalinka
K
Koteva
Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main St W, Hamilton, Ontario, Canada L8S 4K1.
Alba
A
Guarné
Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, Ontario, Canada M5G 1L6.
Alexei
A
Savchenko
M.G. DeGroote Institute for Infectious Disease Research, Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main St W, Hamilton, Ontario, Canada L8S 4K1.
Gerard D
GD
Wright
D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000378:metabolism* / Q000494:pharmacology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D001711:Biotransformation; D018360:Crystallography, X-Ray; D024881:Drug Resistance, Bacterial; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D008089:Listeria monocytogenes / Q000145:classification / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010770:Phosphotransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010802:Phylogeny; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012293:Rifampin / Q000378:metabolism* / Q000494:pharmacology; D016415:Sequence Alignment
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D011994:Recombinant Proteins; D000255:Adenosine Triphosphate; D010770:Phosphotransferases; D012293:Rifampin