Structural Basis for Bisphosphonate-mediated Inhibition of Isoprenoid Biosynthesis

Abstract

Farnesyl pyrophosphate synthetase (FPPS) synthesizes farnesyl pyrophosphate through successive condensations of isopentyl pyrophosphate with dimethylallyl pyrophosphate and geranyl pyrophosphate. Nitrogen-containing bisphosphonate drugs used to treat osteoclast-mediated bone resorption and tumor-induced hypercalcemia are potent inhibitors of the enzyme. Here we present crystal structures of substrate and bisphosphonate complexes of FPPS. The structures reveal how enzyme conformational changes organize conserved active site residues to exploit metal-induced ionization and substrate positioning for catalysis. The structures further demonstrate how nitrogen-containing bisphosphonates mimic a carbocation intermediate to inhibit the enzyme. Together, these FPPS complexes provide a structural template for the design of novel inhibitors that may prove useful for the treatment of osteoporosis and other clinical indications including cancer. Farnesyl pyrophosphate synthetase (FPPS) synthesizes farnesyl pyrophosphate through successive condensations of isopentyl pyrophosphate with dimethylallyl pyrophosphate and geranyl pyrophosphate. Nitrogen-containing bisphosphonate drugs used to treat osteoclast-mediated bone resorption and tumor-induced hypercalcemia are potent inhibitors of the enzyme. Here we present crystal structures of substrate and bisphosphonate complexes of FPPS. The structures reveal how enzyme conformational changes organize conserved active site residues to exploit metal-induced ionization and substrate positioning for catalysis. The structures further demonstrate how nitrogen-containing bisphosphonates mimic a carbocation intermediate to inhibit the enzyme. Together, these FPPS complexes provide a structural template for the design of novel inhibitors that may prove useful for the treatment of osteoporosis and other clinical indications including cancer. Post-translational modification of C-terminal CAAX sequences by covalent attachment of isoprenyl chains is crucial for intracellular localization and proper function of small GTPases such as Ras, Rac, Rho, and CDC42 (1Vicent D. Maratos-Flier E. Kahn C.R. Mol. Cell. Biol. 2000; 20: 2158-2166Crossref PubMed Scopus (23) Google Scholar, 2Holstein S.A. Wohlford-Lenane C.L. Wiemer D.F. Hohl R.J. Biochemistry. 2003; 42: 4384-4391Crossref PubMed Scopus (20) Google Scholar). The substrates for these modifications are the 15-carbon isoprenoid farnesyl pyrophosphate (FPP) 1The abbreviations used are: FPP, farnesyl pyrophosphate; FPPS, farnesyl pyrophosphate synthetase; IPP, isopentyl pyrophosphate; DMAPP, dimethylallyl pyrophosphate; DMSPP, dimethylallyl S-thiolodiphosphate.1The abbreviations used are: FPP, farnesyl pyrophosphate; FPPS, farnesyl pyrophosphate synthetase; IPP, isopentyl pyrophosphate; DMAPP, dimethylallyl pyrophosphate; DMSPP, dimethylallyl S-thiolodiphosphate. or the 20-carbon isoprenoid geranyl-geranyl pyrophosphate synthesized by enzymes of the mevalonate pathway (3Spurgeon S.L. Porter J.W. Porter J.W. Spurgeon S.L. Biosynthesis of Isoprenoid Compounds. John Wiley and Sons, New York1981: 1-93Google Scholar) (Fig. 1A). A key branch point enzyme of the mevalonate pathway is farnesyl pyrophosphate synthetase (FPPS), a ∼30-kDa Mg2+-dependent homodimeric enzyme that synthesizes (E,E)-FPP from isopentyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP) (4Nishino T. Ogura K. Seto S. J. Am. Chem. Soc. 1972; 94: 6849-6853Crossref PubMed Scopus (47) Google Scholar, 5Ding V.D. Sheares B.T. Bergstrom J.D. Ponpipom M.M. Perez L.B. Poulter C.D. Biochem. J. 1991; 275: 61-65Crossref PubMed Scopus (30) Google Scholar) (Fig. 1B).Interest in understanding FPPS activity stems from the recent discovery that FPPS is the molecular target of nitrogen-containing bisphosphonates (6van Beek E. Pieterman E. Cohen L. Lowik C. Papapoulos S. Biochem. Biophys. Res. Commun. 1999; 264: 108-111Crossref PubMed Scopus (465) Google Scholar, 7van Beek E. Pieterman E. Cohen L. Lowik C. Papapoulos S. Biochem. Biophys. Res. Commun. 1999; 255: 491-494Crossref PubMed Scopus (191) Google Scholar, 31Keller R.K. Fliesler S.J. Biochem. Biophys. Res. Commun. 1999; 266: 560-563Crossref PubMed Scopus (84) Google Scholar, 32Bergstrom J.D. Bostedor R.G. Masarachia P.J. Reszka A.A. Rodan G. Arch. Biochem. Biophys. 2000; 373: 231-241Crossref PubMed Scopus (372) Google Scholar). Bisphophonates are non-cleavable pyrophosphate (P-O-P) analogues in which the central oxygen is replaced by a carbon (P-C-P) with various side chains (Fig. 1C). Against parasitic organisms (8Montalvetti A. Fernandez A. Sanders J.M. Ghosh S. Van Brussel E. Oldfield E. Docampo R. J. Biol. Chem. 2003; 278: 17075-17083Abstract Full Text Full Text PDF PubMed Scopus (83) Google Scholar, 9Werbovetz K.A. Expert Opin. Ther. Targets. 2002; 6: 407-422Crossref PubMed Scopus (23) Google Scholar) these agents have been shown in vitro to disrupt cell growth through FPPS inhibition. In people, bisphosphonates are targeted to bone tissue (10Rogers M.J. Curr. Pharm. Des. 2003; 9: 2643-2658Crossref PubMed Scopus (504) Google Scholar) where FPPS inhibition in bone-resorbing osteoclasts is a current therapeutic approach for treating post-menopausal osteoporosis (11Rodan G.A. Reszka A.A. J. Bone Joint Surg. Am. 2003; 85-A: 8-12Crossref PubMed Scopus (21) Google Scholar, 12Watts N.B. Clin. Geriatr. Med. 2003; 19: 395-414Abstract Full Text Full Text PDF PubMed Scopus (81) Google Scholar). Because of their bone-targeting properties, bisphosphonates have also found use as agents to treat tumor-induced hypercalcemia (13Riccardi A. Grasso D. Danova M. Tumori. 2003; 89: 223-236Crossref PubMed Scopus (11) Google Scholar), Paget's disease (14Bartl R. Dtsch. Med. Wochenschr. 2002; 127: 638PubMed Google Scholar), and osteolytic metastases (15Green J.R. Cancer. 2003; 97: 840-847Crossref PubMed Scopus (280) Google Scholar).Although structures of apo- and ligand-bound avian FPPS have been solved (16Tarshis L.C. Yan M. Poulter C.D. Sacchettini J.C. Biochemistry. 1994; 33: 10871-10877Crossref PubMed Scopus (371) Google Scholar, 17Tarshis L.C. Proteau P.J. Kellogg B.A. Sacchettini J.C. Poulter C.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 15018-15023Crossref PubMed Scopus (315) Google Scholar), the active sites are unassembled and do not provide substantial information concerning catalysis. Thus, to resolve the molecular basis of catalysis, and also to understand the structural features governing bisphosphonate recognition, we determined the structures of unliganded Staphylococcus aureus FPPS (FPPS-Sa), as well as two Escherichia coli FPPS (FPPS-Ec) ternary complexes. These ternary complexes include a 2.4-Å “substrate-bound” structure containing and the dimethylallyl Poulter C.D. 2000; PubMed Scopus Google Scholar), and a structure containing and the osteoporosis (Fig. 1C). the E. coli enzyme conserved active site residues with FPPS T. S. M. A. K. M. T. Ogura K. J. Biochem. PubMed Scopus Google Scholar), these complexes provide a structural for the of novel FPPS and FPPS the from E. coli and S. aureus The E. coli to to and to C-terminal the the J.R. J. 9: PubMed Scopus Google Scholar). by and of S. aureus FPPS by of with of D. J. M. D. J. Biol. 2003; PubMed Scopus Google Scholar). and of E. coli FPPS by enzyme with IPP, DMSPP, and or IPP, and and of with of in with and by in from the S. aureus the a of from the E. coli a of and Scopus Google Scholar). of the S. aureus enzyme to the and have two in the of the E. coli enzyme to the and two in the and are in in The structure of S. aureus FPPS determined with the E. G. PubMed Scopus Google Scholar). The E. coli structures determined by molecular J. Biol. PubMed Scopus Google Scholar) with the S. aureus structure as a structures in and and J. Biol. 1999; PubMed Scopus Google aureus structure of the of the and of conformational for enzyme activity (Fig. The is by and and is to the two The and The is by and and are in the of the and the conserved that substrate and catalysis. in avian FPPS (16Tarshis L.C. Yan M. Poulter C.D. Sacchettini J.C. Biochemistry. 1994; 33: 10871-10877Crossref PubMed Scopus (371) Google Scholar, 17Tarshis L.C. Proteau P.J. Kellogg B.A. Sacchettini J.C. Poulter C.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 15018-15023Crossref PubMed Scopus (315) Google Scholar), a and enzyme of conformational changes and the of is the active of the ternary The pyrophosphate is to the active site and is in to FPPS substrate and that the two isoprenoid substrates in a is as FPPS the conserved of and nitrogen-containing for two have been and is as and with from coli FPPS the structural features governing FPPS substrate and we a ternary containing and the FPPS that the from and active site residues in a (Fig. with the unliganded S. aureus and avian structures (16Tarshis L.C. Yan M. Poulter C.D. Sacchettini J.C. Biochemistry. 1994; 33: 10871-10877Crossref PubMed Scopus (371) Google Scholar, 17Tarshis L.C. Proteau P.J. Kellogg B.A. Sacchettini J.C. Poulter C.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 15018-15023Crossref PubMed Scopus (315) Google Scholar), the E. coli ternary complexes reveal conformational changes the enzyme and the and conserved residues that substrate and FPPS to the through a that is by conserved of the (Fig. are by and (Fig. The side of from the two oxygen and The side chains of and from the two and two the and as well as a oxygen and the active site the further the side chains of conserved as well as and These a key in pyrophosphate and substrate of a that the isoprenyl (16Tarshis L.C. Yan M. Poulter C.D. Sacchettini J.C. Biochemistry. 1994; 33: 10871-10877Crossref PubMed Scopus (371) Google Scholar, 17Tarshis L.C. Proteau P.J. Kellogg B.A. Sacchettini J.C. Poulter C.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 15018-15023Crossref PubMed Scopus (315) Google Scholar) (Fig. The to the with residues from and and and of the the of the with and and and the the of the and of is by the of the enzyme that residues the pyrophosphate (Fig. A the C-terminal and side to with two which is also by a C-terminal a to a to the pyrophosphate include with and and with and These pyrophosphate in the FPPS active site such that is conserved and the of (Fig. these the of from the of of the FPPS site and for FPPS isoprenoid the side oxygen of and and the of are with their the The pyrophosphate oxygen that as the to the the of IPP, and the of are substrate ionization by the carbocation that that of in the is by a pyrophosphate oxygen that as the the FPPS that bisphosphonates inhibit FPPS by to the in a to that of (Fig. The conformational with IPP, as well as the to the inhibitors mimic in the substrate The side of the bisphosphonate to the that the of to for DMSPP, as well as a the and conserved the to with have that the by FPPS by a C.D. J.C. J. Biol. Chem. Full Text PDF PubMed Google Scholar) in which the of is by the of the substrate J.W. G. L. J. Biol. Chem. Full Text PDF PubMed Google Scholar). The 2.4-Å structure of the ternary the structural basis for of the FPPS is to substrate with in catalysis. In the substrate and structures with two pyrophosphate a oxygen of the pyrophosphate (Fig. These as well as to the the isoprenoid in a for catalysis. ionization of the by the FPPS carbocation with the and pyrophosphate is the by with the the is by with the pyrophosphate and also through conserved FPPS the oxygen of and the side of and a site by their the the and (Fig. A and substrate that with the of IPP, which is from the (Fig. structure further that a pyrophosphate oxygen is the that the intermediate to the isoprenoid (Fig. A and In the ternary oxygen is by conserved and (Fig. with J.W. G. L. J. Biol. Chem. Full Text PDF PubMed Google Scholar, Poulter C.D. 1999; PubMed Scopus Google Scholar), is to the and from the and that the from these and disrupt the structure of these to pyrophosphate and FPPS bisphosphonates are to inhibit FPPS by the carbocation intermediate substrate ionization Oldfield E. Biochem. Biophys. Res. Commun. 1999; PubMed Scopus Google Scholar). The structure of the FPPS ternary and how key enzyme as well as in to the FPPS by pyrophosphate to the and by the enzyme for nitrogen-containing bisphosphonates is through the side that the site (Fig. DMSPP, the with the and of and may to to the enzyme in recent discovery that bisphosphonates used to treat osteoporosis are potent FPPS inhibitors in understanding the structural features governing inhibition. These FPPS ternary complexes how conformational changes and reveal how bisphosphonates target the active of the enzyme. The complexes the structural basis for bisphosphonate inhibition of isoprenoid and provide a structural template for novel that in treating and parasitic and other Post-translational modification of C-terminal CAAX sequences by covalent attachment of isoprenyl chains is crucial for intracellular localization and proper function of small GTPases such as Ras, Rac, Rho, and CDC42 (1Vicent D. Maratos-Flier E. Kahn C.R. Mol. Cell. Biol. 2000; 20: 2158-2166Crossref PubMed Scopus (23) Google Scholar, 2Holstein S.A. Wohlford-Lenane C.L. Wiemer D.F. Hohl R.J. Biochemistry. 2003; 42: 4384-4391Crossref PubMed Scopus (20) Google Scholar). The substrates for these modifications are the 15-carbon isoprenoid farnesyl pyrophosphate (FPP) 1The abbreviations used are: FPP, farnesyl pyrophosphate; FPPS, farnesyl pyrophosphate synthetase; IPP, isopentyl pyrophosphate; DMAPP, dimethylallyl pyrophosphate; DMSPP, dimethylallyl S-thiolodiphosphate.1The abbreviations used are: FPP, farnesyl pyrophosphate; FPPS, farnesyl pyrophosphate synthetase; IPP, isopentyl pyrophosphate; DMAPP, dimethylallyl pyrophosphate; DMSPP, dimethylallyl S-thiolodiphosphate. or the 20-carbon isoprenoid geranyl-geranyl pyrophosphate synthesized by enzymes of the mevalonate pathway (3Spurgeon S.L. Porter J.W. Porter J.W. Spurgeon S.L. Biosynthesis of Isoprenoid Compounds. John Wiley and Sons, New York1981: 1-93Google Scholar) (Fig. 1A). A key branch point enzyme of the mevalonate pathway is farnesyl pyrophosphate synthetase (FPPS), a ∼30-kDa Mg2+-dependent homodimeric enzyme that synthesizes (E,E)-FPP from isopentyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP) (4Nishino T. Ogura K. Seto S. J. Am. Chem. Soc. 1972; 94: 6849-6853Crossref PubMed Scopus (47) Google Scholar, 5Ding V.D. Sheares B.T. Bergstrom J.D. Ponpipom M.M. Perez L.B. Poulter C.D. Biochem. J. 1991; 275: 61-65Crossref PubMed Scopus (30) Google Scholar) (Fig. in understanding FPPS activity stems from the recent discovery that FPPS is the molecular target of nitrogen-containing bisphosphonates (6van Beek E. Pieterman E. Cohen L. Lowik C. Papapoulos S. Biochem. Biophys. Res. Commun. 1999; 264: 108-111Crossref PubMed Scopus (465) Google Scholar, 7van Beek E. Pieterman E. Cohen L. Lowik C. Papapoulos S. Biochem. Biophys. Res. Commun. 1999; 255: 491-494Crossref PubMed Scopus (191) Google Scholar, 31Keller R.K. Fliesler S.J. Biochem. Biophys. Res. Commun. 1999; 266: 560-563Crossref PubMed Scopus (84) Google Scholar, 32Bergstrom J.D. Bostedor R.G. Masarachia P.J. Reszka A.A. Rodan G. Arch. Biochem. Biophys. 2000; 373: 231-241Crossref PubMed Scopus (372) Google Scholar). Bisphophonates are non-cleavable pyrophosphate (P-O-P) analogues in which the central oxygen is replaced by a carbon (P-C-P) with various side chains (Fig. 1C). Against parasitic organisms (8Montalvetti A. Fernandez A. Sanders J.M. Ghosh S. Van Brussel E. Oldfield E. Docampo R. J. Biol. Chem. 2003; 278: 17075-17083Abstract Full Text Full Text PDF PubMed Scopus (83) Google Scholar, 9Werbovetz K.A. Expert Opin. Ther. Targets. 2002; 6: 407-422Crossref PubMed Scopus (23) Google Scholar) these agents have been shown in vitro to disrupt cell growth through FPPS inhibition. In people, bisphosphonates are targeted to bone tissue (10Rogers M.J. Curr. Pharm. Des. 2003; 9: 2643-2658Crossref PubMed Scopus (504) Google Scholar) where FPPS inhibition in bone-resorbing osteoclasts is a current therapeutic approach for treating post-menopausal osteoporosis (11Rodan G.A. Reszka A.A. J. Bone Joint Surg. Am. 2003; 85-A: 8-12Crossref PubMed Scopus (21) Google Scholar, 12Watts N.B. Clin. Geriatr. Med. 2003; 19: 395-414Abstract Full Text Full Text PDF PubMed Scopus (81) Google Scholar). Because of their bone-targeting properties, bisphosphonates have also found use as agents to treat tumor-induced hypercalcemia (13Riccardi A. Grasso D. Danova M. Tumori. 2003; 89: 223-236Crossref PubMed Scopus (11) Google Scholar), Paget's disease (14Bartl R. Dtsch. Med. Wochenschr. 2002; 127: 638PubMed Google Scholar), and osteolytic metastases (15Green J.R. Cancer. 2003; 97: 840-847Crossref PubMed Scopus (280) Google Scholar). structures of apo- and ligand-bound avian FPPS have been solved (16Tarshis L.C. Yan M. Poulter C.D. Sacchettini J.C. Biochemistry. 1994; 33: 10871-10877Crossref PubMed Scopus (371) Google Scholar, 17Tarshis L.C. Proteau P.J. Kellogg B.A. Sacchettini J.C. Poulter C.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 15018-15023Crossref PubMed Scopus (315) Google Scholar), the active sites are unassembled and do not provide substantial information concerning catalysis. Thus, to resolve the molecular basis of catalysis, and also to understand the structural features governing bisphosphonate recognition, we determined the structures of unliganded Staphylococcus aureus FPPS (FPPS-Sa), as well as two Escherichia coli FPPS (FPPS-Ec) ternary complexes. These ternary complexes include a 2.4-Å “substrate-bound” structure containing and the dimethylallyl Poulter C.D. 2000; PubMed Scopus Google Scholar), and a structure containing and the osteoporosis (Fig. 1C). the E. coli enzyme conserved active site residues with FPPS T. S. M. A. K. M. T. Ogura K. J. Biochem. PubMed Scopus Google Scholar), these complexes provide a structural for the of novel FPPS and FPPS the from E. coli and S. aureus The E. coli to to and to C-terminal the the J.R. J. 9: PubMed Scopus Google Scholar). by and of S. aureus FPPS by of with of D. J. M. D. J. Biol. 2003; PubMed Scopus Google Scholar). and of E. coli FPPS by enzyme with IPP, DMSPP, and or IPP, and and of with of in with and by in from the S. aureus the a of from the E. coli a of and Scopus Google Scholar). of the S. aureus enzyme to the and have two in the of the E. coli enzyme to the and two in the and are in in The structure of S. aureus FPPS determined with the E. G. PubMed Scopus Google Scholar). The E. coli structures determined by molecular J. Biol. PubMed Scopus Google Scholar) with the S. aureus structure as a structures in and and J. Biol. 1999; PubMed Scopus Google Scholar). and FPPS the from E. coli and S. aureus The E. coli to to and to C-terminal the the J.R. J. 9: PubMed Scopus Google Scholar). by and of S. aureus FPPS by of with of D. J. M. D. J. Biol. 2003; PubMed Scopus Google Scholar). and of E. coli FPPS by enzyme with IPP, DMSPP, and or IPP, and and of with of in with and by in from the S. aureus the a of from the E. coli a of and Scopus Google Scholar). of the S. aureus enzyme to the and have two in the of the E. coli enzyme to the and two in the and are in in The structure of S. aureus FPPS determined with the E. G. PubMed Scopus Google Scholar). The E. coli structures determined by molecular J. Biol. PubMed Scopus Google Scholar) with the S. aureus structure as a structures in and and J. Biol. 1999; PubMed Scopus Google Scholar). aureus structure of the of the and of conformational for enzyme activity (Fig. The is by and and is to the two The and The is by and and are in the of the and the conserved that substrate and catalysis. in avian FPPS (16Tarshis L.C. Yan M. Poulter C.D. Sacchettini J.C. Biochemistry. 1994; 33: 10871-10877Crossref PubMed Scopus (371) Google Scholar, 17Tarshis L.C. Proteau P.J. Kellogg B.A. Sacchettini J.C. Poulter C.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 15018-15023Crossref PubMed Scopus (315) Google Scholar), a and enzyme coli FPPS the structural features governing FPPS substrate and we a ternary containing and the FPPS that the from and active site residues in a (Fig. with the unliganded S. aureus and avian structures (16Tarshis L.C. Yan M. Poulter C.D. Sacchettini J.C. Biochemistry. 1994; 33: 10871-10877Crossref PubMed Scopus (371) Google Scholar, 17Tarshis L.C. Proteau P.J. Kellogg B.A. Sacchettini J.C. Poulter C.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 15018-15023Crossref PubMed Scopus (315) Google Scholar), the E. coli ternary complexes reveal conformational changes the enzyme and the and conserved residues that substrate and FPPS to the through a that is by conserved of the (Fig. are by and (Fig. The side of from the two oxygen and The side chains of and from the two and two the and as well as a oxygen and the active site the further the side chains of conserved as well as and These a key in pyrophosphate and substrate of a that the isoprenyl (16Tarshis L.C. Yan M. Poulter C.D. Sacchettini J.C. Biochemistry. 1994; 33: 10871-10877Crossref PubMed Scopus (371) Google Scholar, 17Tarshis L.C. Proteau P.J. Kellogg B.A. Sacchettini J.C. Poulter C.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 15018-15023Crossref PubMed Scopus (315) Google Scholar) (Fig. The to the with residues from and and and of the the of the with and and and the the of the and of is by the of the enzyme that residues the pyrophosphate (Fig. A the C-terminal and side to with two which is also by a C-terminal a to a to the pyrophosphate include with and and with and These pyrophosphate in the FPPS active site such that is conserved and the of (Fig. these the of from the of of the FPPS site and for FPPS isoprenoid the side oxygen of and and the of are with their the The pyrophosphate oxygen that as the to the the of IPP, and the of are substrate ionization by the carbocation that that of in the is by a pyrophosphate oxygen that as the the FPPS that bisphosphonates inhibit FPPS by to the in a to that of (Fig. The conformational with IPP, as well as the to the inhibitors mimic in the substrate The side of the bisphosphonate to the that the of to for DMSPP, as well as a the and conserved the to with S. aureus structure of the of the and of conformational for enzyme activity (Fig. The is by and and is to the two The and The is by and and are in the of the and the conserved that substrate and catalysis. in avian FPPS (16Tarshis L.C. Yan M. Poulter C.D. Sacchettini J.C. Biochemistry. 1994; 33: 10871-10877Crossref PubMed Scopus (371) Google Scholar, 17Tarshis L.C. Proteau P.J. Kellogg B.A. Sacchettini J.C. Poulter C.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 15018-15023Crossref PubMed Scopus (315) Google Scholar), a and enzyme E. coli FPPS the structural features governing FPPS substrate and we a ternary containing and the FPPS that the from and active site residues in a (Fig. with the unliganded S. aureus and avian structures (16Tarshis L.C. Yan M. Poulter C.D. Sacchettini J.C. Biochemistry. 1994; 33: 10871-10877Crossref PubMed Scopus (371) Google Scholar, 17Tarshis L.C. Proteau P.J. Kellogg B.A. Sacchettini J.C. Poulter C.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 15018-15023Crossref PubMed Scopus (315) Google Scholar), the E. coli ternary complexes reveal conformational changes the enzyme and the and conserved residues that substrate and FPPS to the through a that is by conserved of the (Fig. are by and (Fig. The side of from the two oxygen and The side chains of and from the two and two the and as well as a oxygen and the active site the further the side chains of conserved as well as and These a key in pyrophosphate and substrate The of a that the isoprenyl (16Tarshis L.C. Yan M. Poulter C.D. Sacchettini J.C. Biochemistry. 1994; 33: 10871-10877Crossref PubMed Scopus (371) Google Scholar, 17Tarshis L.C. Proteau P.J. Kellogg B.A. Sacchettini J.C. Poulter C.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 15018-15023Crossref PubMed Scopus (315) Google Scholar) (Fig. The to the with residues from and and and of the the of the with and and and the the of the and of is by the of the enzyme that residues the pyrophosphate (Fig. A the C-terminal and side to with two which is also by a C-terminal a to a to the pyrophosphate include with and and with and These pyrophosphate in the FPPS active site such that is conserved and the of (Fig. these the of from the of FPPS FPPS that bisphosphonates inhibit FPPS by to the in a to that of (Fig. The conformational with IPP, as well as the to the inhibitors mimic in the substrate The side of the bisphosphonate to the that the of to for DMSPP, as well as a the and conserved the to with have that the by FPPS by a C.D. J.C. J. Biol. Chem. Full Text PDF PubMed Google Scholar) in which the of is by the of the substrate J.W. G. L. J. Biol. Chem. Full Text PDF PubMed Google Scholar). The 2.4-Å structure of the ternary the structural basis for of the FPPS is to substrate with in catalysis. In the substrate and structures with two pyrophosphate a oxygen of the pyrophosphate (Fig. These as well as to the the isoprenoid in a for catalysis. ionization of the by the FPPS carbocation with the and pyrophosphate is the by with the the is by with the pyrophosphate and also through conserved FPPS the oxygen of and the side of and a site by their the the and (Fig. A and substrate that with the of IPP, which is from the (Fig. structure further that a pyrophosphate oxygen is the that the intermediate to the isoprenoid (Fig. A and In the ternary oxygen is by conserved and (Fig. with J.W. G. L. J. Biol. Chem. Full Text PDF PubMed Google Scholar, Poulter C.D. 1999; PubMed Scopus Google Scholar), is to the and from the and that the from these and disrupt the structure of these to pyrophosphate and FPPS bisphosphonates are to inhibit FPPS by the carbocation intermediate substrate ionization Oldfield E. Biochem. Biophys. Res. Commun. 1999; PubMed Scopus Google Scholar). The structure of the FPPS ternary and how key enzyme as well as in to the FPPS by pyrophosphate to the and by the enzyme for nitrogen-containing bisphosphonates is through the side that the site (Fig. DMSPP, the with the and of and may to to the enzyme in recent discovery that bisphosphonates used to treat osteoporosis are potent FPPS inhibitors in understanding the structural features governing inhibition. These FPPS ternary complexes how conformational changes and reveal how bisphosphonates target the active of the enzyme. The complexes the structural basis for bisphosphonate inhibition of isoprenoid and provide a structural template for novel that in treating and parasitic and other have that the by FPPS by a C.D. J.C. J. Biol. Chem. Full Text PDF PubMed Google Scholar) in which the of is by the of the substrate J.W. G. L. J. Biol. Chem. Full Text PDF PubMed Google Scholar). The 2.4-Å structure of the ternary the structural basis for The of the FPPS is to substrate with in catalysis. In the substrate and structures with two pyrophosphate a oxygen of the pyrophosphate (Fig. These as well as to the the isoprenoid in a for catalysis. ionization of the by the FPPS carbocation with the and pyrophosphate is the by with the the is by with the pyrophosphate and also through conserved FPPS the oxygen of and the side of and a site by their the the and (Fig. A and substrate that with the of IPP, which is from the (Fig. structure further that a pyrophosphate oxygen is the that the intermediate to the isoprenoid (Fig. A and In the ternary oxygen is by conserved and (Fig. with J.W. G. L. J. Biol. Chem. Full Text PDF PubMed Google Scholar, Poulter C.D. 1999; PubMed Scopus Google Scholar), is to the and from the and that the from these and disrupt the structure of these to pyrophosphate and FPPS bisphosphonates are to inhibit FPPS by the carbocation intermediate substrate ionization Oldfield E. Biochem. Biophys. Res. Commun. 1999; PubMed Scopus Google Scholar). The structure of the FPPS ternary and how key enzyme as well as in to the FPPS by pyrophosphate to the and by the enzyme for nitrogen-containing bisphosphonates is through the side that the site (Fig. DMSPP, the with the and of and may to to the enzyme in The recent discovery that bisphosphonates used to treat osteoporosis are potent FPPS inhibitors in understanding the structural features governing inhibition. These FPPS ternary complexes how conformational changes and reveal how bisphosphonates target the active of the enzyme. The complexes the structural basis for bisphosphonate inhibition of isoprenoid and provide a structural template for novel that in treating and parasitic and other G. for D. for and E. and C. for of the and also the which is by the of of of the of the with with