Crystal Structure of Haemophilus influenzae NadR Protein

Abstract

Haemophilus influenzae NadR protein (hiNadR) has been shown to be a bifunctional enzyme possessing both NMN adenylytransferase (NMNAT; EC 2.7.7.1) and ribosylnicotinamide kinase (RNK; EC 2.7.1.22) activities. Its function is essential for the growth and survival of H. influenzae and thus may present a new highly specific anti-infectious drug target. We have solved the crystal structure ofhiNadR complexed with NAD using the selenomethionine MAD phasing method. The structure reveals the presence of two distinct domains. The N-terminal domain that hosts the NMNAT activity is closely related to archaeal NMNAT, whereas the C-terminal domain, which has been experimentally demonstrated to possess ribosylnicotinamide kinase activity, is structurally similar to yeast thymidylate kinase and several other P-loop-containing kinases. There appears to be no cross-talk between the two active sites. The bound NAD at the active site of the NMNAT domain reveals several critical interactions between NAD and the protein. There is also a second non-active-site NAD molecule associated with the C-terminal RNK domain that adopts a highly folded conformation with the nicotinamide ring stacking over the adenine base. Whereas the RNK domain of the hiNadR structure presented here is the first structural characterization of a ribosylnicotinamide kinase from any organism, the NMNAT domain ofhiNadR defines yet another member of the pyridine nucleotide adenylyltransferase family. Haemophilus influenzae NadR protein (hiNadR) has been shown to be a bifunctional enzyme possessing both NMN adenylytransferase (NMNAT; EC 2.7.7.1) and ribosylnicotinamide kinase (RNK; EC 2.7.1.22) activities. Its function is essential for the growth and survival of H. influenzae and thus may present a new highly specific anti-infectious drug target. We have solved the crystal structure ofhiNadR complexed with NAD using the selenomethionine MAD phasing method. The structure reveals the presence of two distinct domains. The N-terminal domain that hosts the NMNAT activity is closely related to archaeal NMNAT, whereas the C-terminal domain, which has been experimentally demonstrated to possess ribosylnicotinamide kinase activity, is structurally similar to yeast thymidylate kinase and several other P-loop-containing kinases. There appears to be no cross-talk between the two active sites. The bound NAD at the active site of the NMNAT domain reveals several critical interactions between NAD and the protein. There is also a second non-active-site NAD molecule associated with the C-terminal RNK domain that adopts a highly folded conformation with the nicotinamide ring stacking over the adenine base. Whereas the RNK domain of the hiNadR structure presented here is the first structural characterization of a ribosylnicotinamide kinase from any organism, the NMNAT domain ofhiNadR defines yet another member of the pyridine nucleotide adenylyltransferase family. nicotinamide mononucleotide nicotinic acid mononucleotide ribosylnicotinamide kinase, NMNAT, NMN adenylyltransferase NaMN adenylyltransferase thymidylate kinase H. influenzae NadR protein pyridine nucleotide adenylyltransferase phosphopantetheine adenylyltransferase Haemophilus influenzae, a small nonmotile Gram-negative bacterium, resides in the upper respiratory mucosa in humans and causes otitis media and respiratory tract infections, mostly in children. This organism lacks almost all of the enzymes necessary for the synthesis of NAD (1Evans N.M. Smith D.D. Wicken A.J. J. Med. Microbiol. 1974; 7: 359-365Crossref PubMed Scopus (65) Google Scholar, 2Fleischmann R.D. Adams M.D. White O. Clayton R.A. Kirkness E.F. Kerlavage A.R. Bult C.J. Tomb J.F. Dougherty B.A. Merrick J.M. McKenney K. Sutton G.G. FitzHugh W. Fields C.A. Gocayne J.D. et al.Science. 1995; 269: 496-512Crossref PubMed Scopus (4702) Google Scholar), and it requires the presence of the so-called V-factors (NADP, NAD, NMN,1 orN-ribosylnicotinamide) in the growth medium (3Gingrich W. Schlenk F. J. Bacteriol. 1944; 47: 535-550Crossref PubMed Google Scholar, 4Shifrine M. Biberstein E.L. Nature. 1960; 187: 623Crossref Scopus (12) Google Scholar). Phosphorylated V-factors (NADP, NAD, and NMN) are degraded by recently identified extracellular and periplasmic hydrolases (5Kemmer G. Reilly T.J. Schmidt-Brauns J. Zlotnik G.W. Green B.A. Fiske M.J. Herbert M. Kraiss A. Schlor S. Smith A. Reidl J. J. Bacteriol. 2001; 183: 3974-3981Crossref PubMed Scopus (66) Google Scholar) toN-ribosylnicotinamide, which is likely to be the only V-factor transported across the inner membrane as an ultimate NAD precursor (6Cynamon M.H. Sorg T.B. Patapow A. J. Gen. Microbiol. 1988; 134: 2789-2799PubMed Google Scholar). Two enzymatic steps are required to convert ribosylnicotinamide to NAD in the cytoplasm: a ribosylnicotinamide kinase (RNK; EC 2.7.1.22) to catalyze the phosphorylation of nicotinamide riboside to produce NMN and a NMN adenylyltransferase (NMNAT; EC 2.7.7.1) to link NMN and the AMP moiety of ATP to generate NAD. Whereas the gene encoding RNK has not been identified in any organism until very recently, 2O. V. Kurnasov, B. M. Polanuyer, S. Ananta, R. Sloutsky, A. Tam, S. Y. Gerdes, A. L. Osterman, submitted for publication. several NMNATs and functionally related NaMNATs (EC. 2.7.7.18) have been characterized at the molecular level from many species during last few years. We will use pyridine nucleotide adenylyltransferase (PNAT) as a generic name for both NMNAT and NaMNAT as well as for enzymes with dual specificities, such as in the case of human NMN/NaMNAT. The three-dimensional structures of PNATs and their complexes with substrate/product have been solved from several sources, includingMethanococcus jannaschii (7D'Angelo I. Raffaelli N. Dabusti V. Lorenzi T. Magni G. Rizzi M. Struct. Fold. Des. 2000; 8: 993-1004Abstract Full Text Full Text PDF Scopus (67) Google Scholar), Methanothermobacter thermautotrophicum (8Saridakis V. Christendat D. Kimber M.S. Dharamsi A. Edwards A.M. Pai E.F. J. Biol. Chem. 2001; 276: 7225-7232Abstract Full Text Full Text PDF PubMed Scopus (49) Google Scholar), Escherichia coli (9Zhang H. Zhou T. Kurnasov O. Cheek S. Grishin N.V. Osterman A. Structure. 2002; 10: 69-79Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar),Bacillus subtilis (10Olland A.M. Underwood K.W. Czerwinski R.M., Lo, M.C. Aulabaugh A. Bard J. Stahl M.L. Somers W.S. Sullivan F.X. Chopra R. J. Biol. Chem. 2002; 277: 3698-3707Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar), and humans (11Garavaglia S. D'Angelo I. Emanuelli M. Carnevali F. Pierella F. Magni G. Rizzi M. J. Biol. Chem. 2002; 277: 8524-8530Abstract Full Text Full Text PDF PubMed Scopus (75) Google Scholar, 12Werner E. Ziegler M. Lerner F. Schweiger M. Heinemann U. FEBS Lett. 2002; 516: 239-244Crossref PubMed Scopus (36) Google Scholar, 13Zhou T. Kurnasov O. Tomchick D.R. Binns D.D. Grishin N.V. Marquez V.E. Osterman A.L. Zhang H. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar). PNATs catalyze the in NAD and encoding have been in almost all with H. influenzae, the of NaMNAT gene is and the function is in the N-terminal domain of the bifunctional NadR which to the E. coli and NadR The NadR the and NMNAT activity has been experimentally in the E. coli enzyme N. Lorenzi T. Emanuelli M. A. S. Magni G. J. Bacteriol. PubMed Google Scholar) recently, in H. influenzae and S. as to the NMNAT activity, NadR in S. and E. a N-terminal to the NMNAT The S. NadR has been demonstrated to be a for the of in both and F. Gen. PubMed Scopus Google Scholar, T. J. Bacteriol. PubMed Google Scholar). also that NadR may with an membrane protein and in the of NAD N. J. Bacteriol. PubMed Google Scholar). H. influenzae NadR lacks the domain, which with the of any of NAD in have and experimentally that RNK activity resides the C-terminal domain of The of NadR for the growth and survival of H. influenzae has also been the crystal structure of H. influenzae NadR protein (hiNadR) complexed with NAD. This structure reveals that whereas the NMNAT domain is mostly similar to the archaeal NMNAT, the C-terminal RNK domain is structurally similar to the yeast thymidylate kinase and several other P-loop-containing nucleotide and kinases. The and of hiNadR protein is the gene encoding of from H. influenzae and a a and site from The the E. coli for The hiNadR protein first with a by a The selenomethionine hiNadR in the in medium with selenomethionine and other S. 276: PubMed Scopus Google Scholar), and using the as the protein. The hiNadR at using the method. hiNadR in and first with of NAD and of ATP and with an of the and the hiNadR to a crystal with a The of to be There is molecule in the to a and at The selenomethionine hiNadR at similar and are of as the at to a all of the and and a at and with the W. 276: PubMed Scopus Google Scholar). The for all are in and of of and are the and structure is the for a of the that from the and are the and structure is the for a of the that from the in a new The of crystal structure solved by the phasing using J. Biol. Chem. PubMed Scopus Google Scholar). of a of and the have a of of with Biol. Chem. 2000; PubMed Scopus Google Scholar), which in a The hiNadR using M. A. 47: PubMed Scopus Google Scholar). The of hiNadR structure using Adams J. M. T. Biol. Chem. PubMed Scopus Google Scholar) with The first of in an of of of and the to a of for the and for for two NAD at the active site of the NMNAT domain and the other associated with the C-terminal RNK domain not in active The for the NAD at the active site of the NMNAT domain from the of NAD in structure (8Saridakis V. Christendat D. Kimber M.S. Dharamsi A. Edwards A.M. Pai E.F. J. Biol. Chem. 2001; 276: 7225-7232Abstract Full Text Full Text PDF PubMed Scopus (49) Google Scholar). The second NAD molecule adopts a highly conformation with the nicotinamide ring stacking over the adenine base. The for NAD molecule to the and conformation with Adams J. M. T. Biol. Chem. PubMed Scopus Google Scholar). also in the of of with in the in a in to to The are in I. with a and using both and in the and for We an with of at for all The at and Two of the protein and in in the with the The hiNadR protein of the of which the NMNAT and RNK domains. We to of the of all NadR has shown that the between hiNadR and other NadR only from the NMNAT of the of hiNadR an N-terminal at in two protein an at This is by the that it is with the of NadR in several other such and The ofhiNadR NMNAT and RNK and is functionally from the The crystal structure of hiNadR and the The first with the N-terminal a and the last at the are in the crystal are two NAD and that are in the ATP present in the no ATP molecule be in the the structure be as The structure of the hiNadR the presence of two distinct by a The N-terminal NMNAT domain adopts a with the the The structure of domain closely archaeal NMNATs (7D'Angelo I. Raffaelli N. Dabusti V. Lorenzi T. Magni G. Rizzi M. Struct. Fold. Des. 2000; 8: 993-1004Abstract Full Text Full Text PDF Scopus (67) Google Scholar, V. Christendat D. Kimber M.S. Dharamsi A. Edwards A.M. Pai E.F. J. Biol. Chem. 2001; 276: 7225-7232Abstract Full Text Full Text PDF PubMed Scopus (49) Google Scholar) with of for with the M. NMNAT The C-terminal RNK domain also adopts a with a of that the in the of the C-terminal RNK domain from that in the NMNAT of the protein structure using L. 1995; Full Text PDF PubMed Scopus Google Scholar) that the RNK domain is structurally similar to several such as yeast thymidylate kinase of of over kinase of of over and kinase of of over of hiNadR with archaeal NMNAT and yeast thymidylate kinase are shown in of hiNadR with archaeal NMNAT and of the of hiNadR NMNAT domain with of the ofhiNadR RNK domain with yeast ofhiNadR with and yeast The and of structural in the is the as in The of NMNAT, the and the of the RNK domain The in that are not are in and the are is only molecule in the of crystal that hiNadR are and the protein is likely to as the of the protein in at two hiNadR and and two and The molecular are and which are the molecular of hiNadR the presence of molecular The at the two protein and two to a a for both of which in the of for the not This that is the species of hiNadR in at The hiNadR in crystal a Two distinct of between are in the and and a that of at the The C-terminal RNK domain from in a between the two of the second with both of the second The other of is between and and and is a of only The has the of a with the between the two of at the of the This a at the of the with a of from and the of the between the The the of the a in the and the of the and from of the to which is at the between in the thus not that several in The active of both NMNAT and RNK are the of the from the NAD associated with the C-terminal RNK of the hiNadR the of the of NAD adopts a highly folded conformation with the nicotinamide ring over the adenine in a The between the adenine and nicotinamide which has been in the as a of of the bound D. PubMed Scopus Google Scholar), is for NAD Whereas of the NAD conformation such as the bound in the active site of NMNAT domain of is only other of NAD that adopts a highly folded the crystal structure of complexed with NAD, the NAD molecule adopts a conformation with a between adenine and nicotinamide of 8: PubMed Scopus Google Scholar). the conformation of the NAD from the second NAD in the structure the of NAD and molecular have in to the NAD in and adopts folded in which the between the nicotinamide and adenine is V. 7: PubMed Scopus Google Scholar, J. Chem. Scopus Google Scholar). There are several specific interactions between the non-active-site NAD molecule The adenine and nicotinamide are between the of and The moiety of the NAD and is in with the of an in the the of the nicotinamide is the of the of and the of the adenine appears to be in with of the that of the non-active-site NAD molecule is a the specific interactions between NAD molecule that it may have E. coli and S. the NadR protein in NAD in a F. Gen. PubMed Scopus Google Scholar, T. J. Bacteriol. PubMed Google Scholar). is that the NAD molecule as an may to a site from the active site of NMNAT of the NAD at site a and to the of NadR to is thus to that the second NAD site in hiNadR may the NAD site in E. coli hiNadR not possess the between the NMNAT and RNK of hiNadR and that of E. and S. NadR are have many structural and of E. coli NadR and S. and structural are to The NAD molecule that in the active site of the N-terminal NMNAT domain of hiNadR adopts an conformation similar to that bound to the (8Saridakis V. Christendat D. Kimber M.S. Dharamsi A. Edwards A.M. Pai E.F. J. Biol. Chem. 2001; 276: 7225-7232Abstract Full Text Full Text PDF PubMed Scopus (49) Google Scholar) The adenine nucleotide site the the between the and the of the first which is also highly the of that the and a and that with the adenine base. molecule in active site is with that in the (8Saridakis V. Christendat D. Kimber M.S. Dharamsi A. Edwards A.M. Pai E.F. J. Biol. Chem. 2001; 276: 7225-7232Abstract Full Text Full Text PDF PubMed Scopus (49) Google Scholar) and appears to the site that be by the and of as shown in the structure of the M. (7D'Angelo I. Raffaelli N. Dabusti V. Lorenzi T. Magni G. Rizzi M. Struct. Fold. Des. 2000; 8: 993-1004Abstract Full Text Full Text PDF Scopus (67) Google Scholar). to the archaeal in hiNadR the to a in the of the nicotinamide of the We the The of the nicotinamide with several from nicotinamide the in to the of whereas the is to the of and which is with in appears to the in a stacking with the nicotinamide the ring of appears to with the nicotinamide ring in a This is in the archaeal NMNAT, two the site and interactions with the nicotinamide the conformation of a between hiNadR and archaeal NMNAT of at the nicotinamide the conformation of the bound NAD from that at the and the nicotinamide ring is at in the two structures The C-terminal RNK domain of hiNadR here is the first structurally characterized RNK from any Its structural to the thymidylate kinase and other it in the and of the nucleotide in T. J. Biol. 1995; PubMed Scopus Google Scholar). 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Full Text Full Text PDF PubMed Scopus Google Scholar). the also in to that in is to the presence of the in the structure the between the two related is not the likely of the it is to at the and the hiNadR the active site of the NMNAT domain is well from that of the RNK domain as by the of the and The between the two active in the is The active of in hiNadR are also from other the two of hiNadR catalyze to be no interactions between the two active the and of the two enzymatic also no such of the of enzymes and at the of NAD and the in the enzymatic and structures of PNATs from are of distinct NAD in several PNATs from species of all of have been characterized and The first is the archaeal NMNAT gene which NMN over NaMN as N. Lorenzi T. Emanuelli M. A. S. Magni G. J. Bacteriol. PubMed Google Scholar). Its structure closely phosphopantetheine adenylyltransferase T. A. J. PubMed Scopus Google Scholar), a and associated and There is also a at the of the protein. The first in the second that with the bound ATP (7D'Angelo I. Raffaelli N. Dabusti V. Lorenzi T. Magni G. Rizzi M. Struct. Fold. Des. 2000; 8: 993-1004Abstract Full Text Full Text PDF Scopus (67) Google Scholar). The of NMNATs are in the bifunctional NMN in M. Carnevali F. F. Pierella F. A. Raffaelli N. Magni G. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google and The N-terminal NMNAT domain of is closely related to archaeal NMNAT both in and and has a for NMN over NaMN N. Lorenzi T. A. Emanuelli M. S. Magni G. FEBS Lett. PubMed Scopus Google Scholar). of the characterized of the second of are the gene in the of a for the NaMN over NMN and be NaMNAT (10Olland A.M. Underwood K.W. Czerwinski R.M., Lo, M.C. Aulabaugh A. Bard J. Stahl M.L. Somers W.S. Sullivan F.X. Chopra R. J. Biol. Chem. 2002; 277: 3698-3707Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar, R.A. Osterman A. 2001; PubMed Google Scholar). The structures of NaMNAT have an with a of (9Zhang H. Zhou T. Kurnasov O. Cheek S. Grishin N.V. Osterman A. Structure. 2002; 10: 69-79Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar, A.M. Underwood K.W. Czerwinski R.M., Lo, M.C. Aulabaugh A. Bard J. Stahl M.L. Somers W.S. Sullivan F.X. Chopra R. J. Biol. Chem. 2002; 277: 3698-3707Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar). The structure of E. coli NaMNAT and reveals that the active site of E. coli NaMNAT is from that of the archaeal NMNAT, and the interactions between protein and the bound NAD a over the pyridine ring (9Zhang H. Zhou T. Kurnasov O. Cheek S. Grishin N.V. Osterman A. Structure. 2002; 10: 69-79Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar). The of for the NaMN that NaMNATs are with the NAD NaMN such as the and a R.A. Osterman A. 2001; PubMed Google Scholar, G. A. Emanuelli M. Raffaelli N. S. Biol. Google Scholar). E. coli NaMNAT is a (9Zhang H. Zhou T. Kurnasov O. Cheek S. Grishin N.V. Osterman A. Structure. 2002; 10: 69-79Abstract Full Text Full Text PDF PubMed Scopus (63) Google subtilis NaMNAT shown to be a (10Olland A.M. Underwood K.W. Czerwinski R.M., Lo, M.C. Aulabaugh A. Bard J. Stahl M.L. Somers W.S. Sullivan F.X. Chopra R. J. Biol. Chem. 2002; 277: 3698-3707Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar), that are in the The all of the identified by the human enzymes have dual and both NMN and NaMN almost well G. A. Emanuelli M. Raffaelli N. S. Biol. Google M. Raffaelli N. S. A. Magni G. PubMed Scopus Google Scholar). are of in of NAD both NMN and NaMN T. Kurnasov O. Tomchick D.R. Binns D.D. Grishin N.V. Marquez V.E. Osterman A.L. Zhang H. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar). The structures of complexed with NAD and with that an active site molecule appears to in the dual T. Kurnasov O. Tomchick D.R. Binns D.D. Grishin N.V. Marquez V.E. Osterman A.L. Zhang H. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar). shown to be a in the crystal structures (11Garavaglia S. D'Angelo I. Emanuelli M. Carnevali F. Pierella F. Magni G. Rizzi M. J. Biol. Chem. 2002; 277: 8524-8530Abstract Full Text Full Text PDF PubMed Scopus (75) Google Scholar, 12Werner E. Ziegler M. Lerner F. Schweiger M. Heinemann U. FEBS Lett. 2002; 516: 239-244Crossref PubMed Scopus (36) Google Scholar, 13Zhou T. Kurnasov O. Tomchick D.R. Binns D.D. Grishin N.V. Marquez V.E. Osterman A.L. Zhang H. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar). that molecular species likely with the in T. Kurnasov O. Tomchick D.R. Binns D.D. Grishin N.V. Marquez V.E. Osterman A.L. Zhang H. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar). the of appears to be similar to that of B. subtilis NaMNAT (10Olland A.M. Underwood K.W. Czerwinski R.M., Lo, M.C. Aulabaugh A. Bard J. Stahl M.L. Somers W.S. Sullivan F.X. Chopra R. J. Biol. Chem. 2002; 277: 3698-3707Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar, 13Zhou T. Kurnasov O. Tomchick D.R. Binns D.D. Grishin N.V. Marquez V.E. Osterman A.L. Zhang H. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar). The NMNAT domain of the NadR yet another of with C-terminal RNK domain, NadR protein convert the ribosylnicotinamide to NAD, thus a critical in the of the NAD have shown that NadR NMN over NaMN N. Lorenzi T. Emanuelli M. A. S. Magni G. J. Bacteriol. PubMed Google with the structures of have the structural for the enzymatic of enzymes be structures also to a of all identified and to with the related phosphopantetheine and for the of between protein recently V. and N. V. in Scholar) to to their This the to between and the in of to the between two with the acid as the and as a in a in such a that the the between the by the function V. and N. V. in Scholar). is from the of PNATs in the that two well archaeal NMNAT gene and NadR and NaMNAT gene and the of two of PNATs are distinct in enzymatic and active site structures are for the the NMNAT from M. jannaschii (7D'Angelo I. Raffaelli N. Dabusti V. Lorenzi T. Magni G. Rizzi M. Struct. Fold. Des. 2000; 8: 993-1004Abstract Full Text Full Text PDF Scopus (67) Google Scholar), M. (8Saridakis V. Christendat D. Kimber M.S. Dharamsi A. Edwards A.M. Pai E.F. J. Biol. Chem. 2001; 276: 7225-7232Abstract Full Text Full Text PDF PubMed Scopus (49) Google Scholar), and the NadR from H. influenzae presented in the all of a nicotinamide between and of the interactions with the nicotinamide of the interactions with the and the stacking of a with the pyridine ring all from nicotinamide structures are for the the NaMNAT coli (9Zhang H. Zhou T. Kurnasov O. Cheek S. Grishin N.V. Osterman A. Structure. 2002; 10: 69-79Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar), B. subtilis (10Olland A.M. Underwood K.W. Czerwinski R.M., Lo, M.C. Aulabaugh A. Bard J. Stahl M.L. Somers W.S. Sullivan F.X. Chopra R. J. Biol. Chem. 2002; 277: 3698-3707Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar), and human (11Garavaglia S. D'Angelo I. Emanuelli M. Carnevali F. Pierella F. Magni G. Rizzi M. J. Biol. Chem. 2002; 277: 8524-8530Abstract Full Text Full Text PDF PubMed Scopus (75) Google Scholar, 12Werner E. Ziegler M. Lerner F. Schweiger M. Heinemann U. FEBS Lett. 2002; 516: 239-244Crossref PubMed Scopus (36) Google Scholar, 13Zhou T. Kurnasov O. Tomchick D.R. Binns D.D. Grishin N.V. Marquez V.E. Osterman A.L. Zhang H. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar). two and and the other between and and the of are in the of nicotinic The critical that a stacking with the pyridine ring from the second and the with from both T. Kurnasov O. Tomchick D.R. Binns D.D. Grishin N.V. Marquez V.E. Osterman A.L. Zhang H. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar). has been that the appears closely related to phosphopantetheine to the PNATs (9Zhang H. Zhou T. Kurnasov O. Cheek S. Grishin N.V. Osterman A. Structure. 2002; 10: 69-79Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar). also reveals that are two distinct of phosphopantetheine phosphopantetheine by the gene that are to and the recently identified phosphopantetheine M. B. M. M. A. V. Osterman A. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar) that are to to their H. influenzae, the NadR protein the only activity in the The RNK activity in NadR also shown to be of for the NAD in H. influenzae (6Cynamon M.H. Sorg T.B. Patapow A. J. Gen. Microbiol. 1988; 134: 2789-2799PubMed Google Scholar, M.H. PubMed Scopus Google Scholar). The of hiNadR for the growth and survival of H. influenzae has recently been and may present a anti-infectious drug specific for H. influenzae and several other The structural that the active site and interactions ofhiNadR will be critical for the and of for essential protein. We and Heinemann for the human NMNAT and Binns for with the and and Tomchick for with and