Structural basis for phosphatidylinositol-phosphate biosynthesis.
Phosphatidylinositol is critical for intracellular signalling and anchoring of carbohydrates and proteins to outer cellular membranes. The defining step in phosphatidylinositol biosynthesis is catalysed by CDP-alcohol phosphotransferases, transmembrane enzymes that use CDP-diacylglycerol as donor substrate for this reaction, and either inositol in eukaryotes or inositol phosphate in prokaryotes as the acceptor alcohol. Here we report the structures of a related enzyme, the phosphatidylinositol-phosphate synthase from Renibacterium salmoninarum, with and without bound CDP-diacylglycerol to 3.6 and 2.5 Å resolution, respectively. These structures reveal the location of the acceptor site, and the molecular determinants of substrate specificity and catalysis. Functional characterization of the 40%-identical ortholog from Mycobacterium tuberculosis, a potential target for the development of novel anti-tuberculosis drugs, supports the proposed mechanism of substrate binding and catalysis. This work therefore provides a structural and functional framework to understand the mechanism of phosphatidylinositol-phosphate biosynthesis.
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA.
Oliver B
OB
Clarke
Department of Physiology and Cellular Biophysics, Columbia University, New York, NY 10032, USA.
David
D
Tomasek
Biology Division, Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Avenida da República-EAN, 2780-157 Oeiras, Portugal.
Carla D
CD
Jorge
Department of Physiology and Cellular Biophysics, Columbia University, New York, NY 10032, USA.
Meagan Belcher
MB
Dufrisne
Department of Physiology and Cellular Biophysics, Columbia University, New York, NY 10032, USA.
Minah
M
Kim
NE-CAT and Department of Chemistry and Chemical Biology, Cornell University, Argonne National Laboratory, Argonne, IL 60439, USA.
Surajit
S
Banerjee
NE-CAT and Department of Chemistry and Chemical Biology, Cornell University, Argonne National Laboratory, Argonne, IL 60439, USA.
Kanagalaghatta R
KR
Rajashankar
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA.
Lawrence
L
Shapiro
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA.
Wayne A
WA
Hendrickson
Biology Division, Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Avenida da República-EAN, 2780-157 Oeiras, Portugal.
Helena
H
Santos
Department of Physiology and Cellular Biophysics, Columbia University, New York, NY 10032, USA.
Filippo
F
Mancia
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051607:CDP-Diacylglycerol-Inositol 3-Phosphatidyltransferase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018360:Crystallography, X-Ray; D007700:Kinetics; D008835:Micrococcaceae / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000737:chemistry / Q000201:enzymology; D018129:Phosphatidylinositol Phosphates / Q000096:biosynthesis*
D001426:Bacterial Proteins; D018129:Phosphatidylinositol Phosphates; D051607:CDP-Diacylglycerol-Inositol 3-Phosphatidyltransferase