Allosteric Activation by Dimerization of the PknD Receptor Ser/Thr Protein Kinase from Mycobacterium tuberculosis

Abstract

To define how extracellular signals activate bacterial receptor Ser/Thr protein kinases, we characterized the regulatory functions of a weak dimer interface identified in the Mycobacterium tuberculosis PknB and PknE receptor kinases. Sequence comparisons revealed that the analogous interface is conserved in PknD orthologs from diverse bacterial species. To analyze the roles of dimerization, we constructed M. tuberculosis PknD kinase domain (KD) fusion proteins that formed dimers upon addition of rapamycin. Dimerization of unphosphorylated M. tuberculosis PknD KD fusions stimulated phosphorylation activity. Mutations in the dimer interface reduced this activation, limited autophosphorylation, and altered substrate specificity. In contrast, an inactive catalytic site mutant retained the ability to stimulate the wild-type KD by dimerization. These results support the idea that dimer formation allosterically activates unphosphorylated PknD. The phosphorylated PknD KD was fully active even in the absence of dimerization, suggesting that phosphorylation provides an additional regulatory mechanism. The conservation of analogous dimers in diverse prokaryotic and eukaryotic Ser/Thr protein kinases implies that this mechanism of protein kinase regulation is ancient and broadly distributed. To define how extracellular signals activate bacterial receptor Ser/Thr protein kinases, we characterized the regulatory functions of a weak dimer interface identified in the Mycobacterium tuberculosis PknB and PknE receptor kinases. Sequence comparisons revealed that the analogous interface is conserved in PknD orthologs from diverse bacterial species. To analyze the roles of dimerization, we constructed M. tuberculosis PknD kinase domain (KD) fusion proteins that formed dimers upon addition of rapamycin. Dimerization of unphosphorylated M. tuberculosis PknD KD fusions stimulated phosphorylation activity. Mutations in the dimer interface reduced this activation, limited autophosphorylation, and altered substrate specificity. In contrast, an inactive catalytic site mutant retained the ability to stimulate the wild-type KD by dimerization. These results support the idea that dimer formation allosterically activates unphosphorylated PknD. The phosphorylated PknD KD was fully active even in the absence of dimerization, suggesting that phosphorylation provides an additional regulatory mechanism. The conservation of analogous dimers in diverse prokaryotic and eukaryotic Ser/Thr protein kinases implies that this mechanism of protein kinase regulation is ancient and broadly distributed. Tight regulation of Ser/Thr protein kinases (STPKs) 2The abbreviations used are: STPK, Ser/Thr protein kinase; EGF, epidermal growth factor; EGFR, EGF receptor; FKBP, FK-506-binding protein; FRAP, FK506-binding protein-rapamycin-associated protein; KD, kinase domain; MyBP, myelin basic protein; PKR, dsRNA-dependent protein kinase R; TCEP, Tris(2-carboxyethyl)phosphine hydrochloride; WT, wild-type; BisTris, 2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diol; dsRNA, double-stranded RNA. 2The abbreviations used are: STPK, Ser/Thr protein kinase; EGF, epidermal growth factor; EGFR, EGF receptor; FKBP, FK-506-binding protein; FRAP, FK506-binding protein-rapamycin-associated protein; KD, kinase domain; MyBP, myelin basic protein; PKR, dsRNA-dependent protein kinase R; TCEP, Tris(2-carboxyethyl)phosphine hydrochloride; WT, wild-type; BisTris, 2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diol; dsRNA, double-stranded RNA. is essential for signaling in all three kingdoms of life. In keeping with the importance of phospho-signaling pathways, STPKs are subject to multiple mechanisms of allosteric regulation (1Huse M. Kuriyan J. Cell. 2002; 109: 275-282Abstract Full Text Full Text PDF PubMed Scopus (1353) Google Scholar). Diverse regulatory surfaces of kinase domains (KDs) can bind additional proteins (as in the cyclin-dependent kinases and cAMP-dependent protein kinase) or other domains of the kinase itself (such as the Src homology 2 and linker domains of Src). These interactions often affect the assembly of the catalytic site by positioning a conserved, substrate-binding element called the C helix. In addition, phosphorylation of a conserved motif called the activation loop often relieves a steric blockade of the active site and promotes assembly of a substrate-binding platform (1Huse M. Kuriyan J. Cell. 2002; 109: 275-282Abstract Full Text Full Text PDF PubMed Scopus (1353) Google Scholar). Although many regulatory interactions have been characterized in eukaryotic protein kinases, the mechanisms by which environmental signals regulate prokaryotic STPKs have yet to be elucidated. Prokaryotic STPKs occur in numerous pathogens and organisms with complex developmental pathways (2Greenstein A.E. Grundner C. Echols N. Gay L.M. Lombana T.N. Miecskowski C.A. Pullen K.E. Sung P.Y. Alber T. J. Mol. Microbiol. Biotechnol. 2005; 9: 167-181Crossref PubMed Scopus (82) Google Scholar). Like their eukaryotic homologs, the bacterial STPKs adopt a characteristic two-domain fold with the ATP-binding site located between the N- and C-terminal lobes (3Gay L.M. Ng H.L. Alber T. J. Mol. Biol. 2006; 360: 409-420Crossref PubMed Scopus (51) Google Scholar, 4Ortiz-Lombardia M. Pompeo F. Boitel B. Alzari P.M. J. Biol. Chem. 2003; 278: 13094-13100Abstract Full Text Full Text PDF PubMed Scopus (131) Google Scholar, 5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar). Unexpectedly, the catalytic domain of the receptor STPK PknB from Mycobacterium tuberculosis formed an unstable, back-to-back dimer through an interface in the N-terminal lobe (Fig. 1A) (3Gay L.M. Ng H.L. Alber T. J. Mol. Biol. 2006; 360: 409-420Crossref PubMed Scopus (51) Google Scholar, 4Ortiz-Lombardia M. Pompeo F. Boitel B. Alzari P.M. J. Biol. Chem. 2003; 278: 13094-13100Abstract Full Text Full Text PDF PubMed Scopus (131) Google Scholar, 5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar). Residues in the PknB dimer interface were found to be strictly conserved in orthologs from dozens of bacterial genera, supporting the proposal that KD dimerization plays an important functional role (5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar). Because the dimer interface included contacts near the C terminus of the C helix, it was proposed that dimerization regulates PknB (5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar). This idea was reinforced by the structure of the M. tuberculosis PknD extracellular domain, which includes a rigid β-propeller motif flexibly tethered to the single, predicted transmembrane helix (6Good M.C. Greenstein A.E. Young T.A. Ng H.L. Alber T. J. Mol. Biol. 2004; 339: 459-469Crossref PubMed Scopus (60) Google Scholar). This arrangement was consistent with a model in which dimerization or localization of the extracellular domain, rather than a conformational change transmitted across the membrane, serves to regulate the intracellular kinase domains (2Greenstein A.E. Grundner C. Echols N. Gay L.M. Lombana T.N. Miecskowski C.A. Pullen K.E. Sung P.Y. Alber T. J. Mol. Microbiol. Biotechnol. 2005; 9: 167-181Crossref PubMed Scopus (82) Google Scholar, 5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar, 6Good M.C. Greenstein A.E. Young T.A. Ng H.L. Alber T. J. Mol. Biol. 2004; 339: 459-469Crossref PubMed Scopus (60) Google Scholar). Further support of a regulatory role for dimerization was provided by the crystal structure of the M. tuberculosis PknE KD, which crystallized as a back-to-back dimer similar to that formed by the PknB KD (Fig. 1A), despite the absence of sequence homology in the interfacial residues (3Gay L.M. Ng H.L. Alber T. J. Mol. Biol. 2006; 360: 409-420Crossref PubMed Scopus (51) Google Scholar). Sequence analysis of the 11 M. tuberculosis STPKs indicated that the predicted dimer interface residues are more similar in PknB, -A, and -L; PknD, -E, and -H; and PknF, -I, and -J, but these three clades are distinct from each other (Fig. 1B) (3Gay L.M. Ng H.L. Alber T. J. Mol. Biol. 2006; 360: 409-420Crossref PubMed Scopus (51) Google Scholar). This conservation within three paralogous branches of M. tuberculosis STPKs supported the importance of the back-to-back pairing interface. Consistent with the proposed regulatory role in PknB and PknE, formation of a structurally similar dimer was found to activate the human dsRNA-dependent protein kinase R (PKR; Fig. 1A) KD in response to double-stranded RNA binding to the ligand-binding domain (7Dar A.C. Dever T.E. Sicheri F. Cell. 2005; 122: 887-900Abstract Full Text Full Text PDF PubMed Scopus (287) Google Scholar, 8Dey M. Cao C. Dar A.C. Tamura T. Ozato K. Sicheri F. Dever T.E. Cell. 2005; 122: 901-913Abstract Full Text Full Text PDF PubMed Scopus (267) Google Scholar). Here we provide experimental evidence that dimerization activates M. tuberculosis PknD. To test the functional roles of dimerization, we used rapamycin to bring together FKBP (FK506-binding protein) and FRAP (FK506-binding proteinrapamycin-associated protein) fusions to the PknD KD. Dimerization strongly activated the unphosphorylated PknD KD, particularly when tested using a cognate in vivo substrate. In contrast, dimerization caused little or no further activation of the active, autophosphorylated KD fusions. The predicted intersubunit contact residues were found to be conserved in orthologs of M. tuberculosis PknD. Mutation of conserved residues in the predicted dimer interface reduced activation, inhibited autophosphorylation, and altered substrate specificity. Remarkably, association with a catalytically inactive mutant containing a normal dimer interface activated the unphosphorylated wild-type (WT) KD fusion. These data support the conclusion that back-to-back dimerization allosterically regulates diverse protein kinases in prokaryotes and eukaryotes. Sequence Comparisons—Ser/Thr kinase sequences were extracted from the NCBI RefSeq data base (9Pruitt K.D. Tatusova T. Maglott D.R. Nucleic Acids Res. 2005; 33: D501-D504Crossref PubMed Scopus (1302) Google Scholar) using BLAST searches (10Altschul S.F. Madden T.L. Schaffer A.A. Zhang J. Zhang Z. Miller W. Lipman D.J. Nucleic Acids Res. 1997; 25: 3389-3402Crossref PubMed Scopus (59732) Google Scholar) against PknB, PknE, and representative eukaryotic kinases. Domain assignments were made with hmmpfam using the PFAM data base (11Finn R.D. Mistry J. Schuster-Bockler B. Griffiths-Jones S. Hollich V. Lassmann T. Moxon S. Marshall M. Khanna A. Durbin R. Eddy S.R. Sonnhammer E.L. Bateman A. Nucleic Acids Res. 2006; PubMed Scopus Google Scholar). containing or domains were and the kinase domains were using Nucleic Acids Res. 2004; PubMed Scopus Google Scholar). M. F. T. N. Nucleic Acids Res. 2005; 33: PubMed Scopus Google Scholar) was used to the sequence conservation the of PknB or was by 2 between the and to the of FKBP and the functional domain of FRAP J. J. J. PubMed Scopus Google Scholar) were from a provided by of with a site and a FKBP or FRAP was with and and to the and In M. tuberculosis PknD, the KD is by an linker called the a transmembrane helix, a and a β-propeller to the of PknD was with and from M. tuberculosis as (5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar). The PknD was or to or and were as (5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar). were to an of in to for using and for an additional The were by and in and Tris(2-carboxyethyl)phosphine were by and the was by The was a with and in with and TCEP, each protein was further by using a To the the was from PknD by with of in and of was with K.E. Ng H.L. Sung P.Y. M.C. Alber T. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar) and with The and were using and the absence of was by of an were in TCEP, and 2 KD fusion protein was to a of basic protein and N-terminal and protein A.E. J.A. Falick A.M. Alber T. Google Scholar) were and rapamycin of in or an of was to each This was for and the was by of in 2 The was were with of were by and of to the from of was were by The were and was with a The and were and phosphorylation were and to the of were three The of dimerization were using a activation model by the of the signals each in the and absence of rapamycin. PknD was and as for the The protein was TCEP, and a in the and with a in The protein was to The of the protein was by The mutant was further by to the The of the as an in vivo substrate of PknD A.E. J.A. Falick A.M. Alber T. Google Scholar) an to the and of an M. tuberculosis STPK using a cognate substrate. of binding to and of in vivo A.E. J.A. Falick A.M. Alber T. Google Scholar). other M. tuberculosis STPK (2Greenstein A.E. Grundner C. Echols N. Gay L.M. Lombana T.N. Miecskowski C.A. Pullen K.E. Sung P.Y. Alber T. J. Mol. Microbiol. Biotechnol. 2005; 9: 167-181Crossref PubMed Scopus (82) Google Scholar, V. K. S. A. 2006; PubMed Scopus Google Scholar, 2005; PubMed Scopus Google Scholar, V. C. 2003; PubMed Scopus Google Scholar, J. R. J. Res. 2006; PubMed Scopus Google Scholar, K. M. A. N. J. 2006; PubMed Scopus Google Scholar, A. R. A. C. Alzari P.M. J. Mol. Biol. 2005; PubMed Scopus Google a domain and is phosphorylated a PknD the role of dimerization in M. tuberculosis PknD, we the conservation of predicted residues in the of PknD The PknD KD is and to the of PknB and PknE, but the structure and for dimerization of the PknD KD tethered to the are PFAM domain assignments for all prokaryotic kinase sequences in the NCBI RefSeq data base identified PknD orthologs with domain These receptor kinases a KD through a transmembrane helix to extracellular predicted to a The sequence with M. tuberculosis PknD. the of sequence conservation the crystal structure of the KD of M. tuberculosis PknE revealed that the predicted dimer interface of PknD is conserved in orthologs (Fig. In with the conservation of the dimer interface in PknB orthologs (5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google this analysis that dimerization is conserved in prokaryotic receptor of PknD test the role of the dimer interface in PknD we the of dimerization kinase activity. Because the M. tuberculosis are used in (3Gay L.M. Ng H.L. Alber T. J. Mol. Biol. 2006; 360: 409-420Crossref PubMed Scopus (51) Google N. Lombana and T. we the PknD KD to proteins that in the of the rapamycin. The fusions the of PknD, the kinase domain and the linker This intracellular of PknD was to the FKBP or the binding domain of the FRAP FKBP rapamycin with and FRAP the complex J. J. J. PubMed Scopus Google Scholar). This binding together the PknD with similar to that be by dimerization of the extracellular domain of the receptor (Fig. dimerization the of the and formation of KD using a of PknD KD fusions that were phosphorylated in that rapamycin little or no in kinase (Fig. was three as the KD a substrate with multiple phosphorylation myelin basic protein and the in vivo PknD dimerization of the phosphorylated PknD KD phosphorylation of MyBP, and it little or no or the PknD KD itself (Fig. important in these is the of this KD in three in the activation were in the PknD KD and R. A. M. A. Boitel B. Alzari P.M. C. Res. 2005; PubMed Scopus Google Scholar). Because phosphorylation is for kinases, PknB (1Huse M. Kuriyan J. Cell. 2002; 109: 275-282Abstract Full Text Full Text PDF PubMed Scopus (1353) Google Scholar, 5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar, R. A. M. A. Boitel B. Alzari P.M. C. Res. 2005; PubMed Scopus Google Scholar, B. M. R. Pompeo F. C. Alzari P.M. Mol. Microbiol. 2003; PubMed Scopus Google the of the KD be to stimulate the the of an in upon dimerization the of the of phosphorylation in the KD. To test the idea that PknD the of dimerization, we the of dimerization the activation of the unphosphorylated PknD KD. The KD fusions to FKBP and FRAP were with the M. tuberculosis Ser/Thr to all the the proteins K.E. Ng H.L. Sung P.Y. M.C. Alber T. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, R. A. M. A. Boitel B. Alzari P.M. C. Res. 2005; PubMed Scopus Google Scholar). The to as by were from the KD fusion reduced the of the PknD KD to (Fig. Because the phosphorylated KD from the of the the from autophosphorylated KD. This an for the of the unphosphorylated PknD KD. dimerization of the unphosphorylated KD stimulated and phosphorylation (Fig. the cognate M. tuberculosis substrate a in was when the KD was with rapamycin. In contrast, rapamycin no the of each fusion protein in (Fig. when the FKBP and FRAP fusions were rapamycin dimerization and These results that dimerization strongly phosphorylation of cognate and phosphorylation of a substrate. these data that dimerization further activate autophosphorylated kinase but rather the of KD. and the role of back-to-back contacts in kinase activation, we predicted residues to and the kinase contacts across the dimer interface in the PknE and is conserved in the predicted dimer interface of all PknD orthologs (Fig. revealed that of the protein from was reduced by the and by (Fig. and Because activates PknB and other protein kinases (1Huse M. Kuriyan J. Cell. 2002; 109: 275-282Abstract Full Text Full Text PDF PubMed Scopus (1353) Google Scholar, 5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar, R. A. M. A. Boitel B. Alzari P.M. C. Res. 2005; PubMed Scopus Google Scholar, B. M. R. Pompeo F. C. Alzari P.M. Mol. Microbiol. 2003; PubMed Scopus Google the of the PknD KD and comparisons of the of these PknD of R. A. M. A. Boitel B. Alzari P.M. C. Res. 2005; PubMed Scopus Google to (11Finn R.D. Mistry J. Schuster-Bockler B. Griffiths-Jones S. Hollich V. Lassmann T. Moxon S. Marshall M. Khanna A. Durbin R. Eddy S.R. Sonnhammer E.L. Bateman A. Nucleic Acids Res. 2006; PubMed Scopus Google M. Cao C. Dar A.C. Tamura T. Ozato K. Sicheri F. Dever T.E. Cell. 2005; 122: 901-913Abstract Full Text Full Text PDF PubMed Scopus (267) Google in a used to this we the of dimerization the of KD fusions as from with were using the phosphorylated WT, and KD fusions from (Fig. The mutant was inactive all in which kinase was The mutant phosphorylated the KD MyBP, and (Fig. Like the protein (Fig. dimerization the of the autophosphorylated KD. In addition, we tested the of each mutant kinase The mutant was inactive in all tested of by the mutant was in the or absence of rapamycin (Fig. in contrast, was stimulated by dimerization (Fig. These results that dimerization activate the KD to cognate of KD of PknD by a allosteric transmitted through the dimer interface are to activate PknD, we a of PknD with an mutant that the kinase catalytically This KD mutant was active than A.E. J.A. Falick A.M. Alber T. Google but the protein was and in the FKBP fusion of the mutant no was with the FRAP fusion of the dimerization activated the KD in a analogous to the KD (Fig. and of and in response to dimerization. to the unphosphorylated KD, dimerization caused the in the of In contrast, a of the FKBP fusions are together by of and no by rapamycin (Fig. These results that a mutant kinase activated the KD by dimerization. together with sequence comparisons (3Gay L.M. Ng H.L. Alber T. J. Mol. Biol. 2006; 360: 409-420Crossref PubMed Scopus (51) Google Scholar, 5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar) (Fig. 1B) and analogous dimers of PknB and PknE KD (3Gay L.M. Ng H.L. Alber T. J. Mol. Biol. 2006; 360: 409-420Crossref PubMed Scopus (51) Google Scholar, 4Ortiz-Lombardia M. Pompeo F. Boitel B. Alzari P.M. J. Biol. Chem. 2003; 278: 13094-13100Abstract Full Text Full Text PDF PubMed Scopus (131) Google Scholar, 5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar) (Fig. 1A), these data support the idea that dimerization through the conserved, back-to-back interface PknD and specificity. of the cognate by the unphosphorylated KD the by dimer The and or inhibited the activation of the PknD KD, suggesting that activation formation of a structurally These define the of an allosteric activation in PknD, which includes the other predicted residues conserved in PknD orthologs (Fig. The of dimerization to stimulate and phosphorylation in the and (Fig. that activation of the KD results from the formation of intersubunit the of in the The back-to-back arrangement of the in the which the active of the from each other (Fig. 1A), the idea that dimerization allosterically activates the KD, rather than results support a model of PknD activation in which kinase the inactive (Fig. Dimerization of the extracellular domains together the intracellular which promotes back-to-back dimerization and activates the kinase for and The that dimerization activated the KD (Fig. and the catalytically inactive KD activated the KD (Fig. provide support for this allosteric mechanism. The of the that KD dimer formation is for with other STPKs (2Greenstein A.E. Grundner C. Echols N. Gay L.M. Lombana T.N. Miecskowski C.A. Pullen K.E. Sung P.Y. Alber T. J. Mol. Microbiol. Biotechnol. 2005; 9: 167-181Crossref PubMed Scopus (82) Google Scholar, R. A. M. A. Boitel B. Alzari P.M. C. Res. 2005; PubMed Scopus Google Scholar, B. M. R. Pompeo F. C. Alzari P.M. Mol. Microbiol. 2003; PubMed Scopus Google activated the PknD KD (Fig. little or no further activation upon dimerization of the autophosphorylated KD (Fig. suggesting that the phosphorylated be active the are dimer formation be important for activity. phosphorylation provides an additional mechanism to activate the PknD KD. data support a model in which of the and are to the phosphorylated kinase to the inactive Consistent with the that a catalytically inactive mutant KD activates in (Fig. of PknD in M. tuberculosis phosphorylation of proteins in vivo A.E. J.A. Falick A.M. Alber T. Google Scholar). These proteins are phosphorylated upon of are to PknD itself as as a of This in phosphorylation upon of a catalytically inactive kinase be an of protein or a of the of other The of the be by activation of the PknD by dimerization with the of inactive mutant in The role of dimerization in PknD the of how this mechanism The conservation of the dimer interface in orthologs of PknD and PknB (5Young T.A. Delagoutte B. Endrizzi J.A. Falick A.M. Alber T. Nat. Struct. Biol. 2003; 10: 168-174Crossref PubMed Scopus (196) Google Scholar) and the conservation of the back-to-back dimer in PknE (3Gay L.M. Ng H.L. Alber T. J. Mol. Biol. 2006; 360: 409-420Crossref PubMed Scopus (51) Google Scholar) support the idea that dimerization regulates these kinases in numerous bacterial Consistent with the allosteric M. tuberculosis PknB, PknD, PknE, and all domains to the KD M. Microbiol. Full Text Full Text PDF PubMed Scopus Google Scholar). a the transmembrane and ligand-binding domains of the B. PknB was to dimers in vivo A. A. M. S. Mol. Microbiol. 2002; PubMed Scopus Google Scholar). Although of the M. tuberculosis have weak (3Gay L.M. Ng H.L. Alber T. J. Mol. Biol. 2006; 360: 409-420Crossref PubMed Scopus (51) Google N. Lombana and T. this dimer is for in which is and in which signaling dimerization. The dimerization mechanism (Fig. is to the activation of all the M. tuberculosis of these proteins a domain with a fold or The predicted extracellular domains of PknF, and sequences with and predicted to occur the Although it is that binding proteins these extracellular these kinases be by by activated STPK or by dimerization that by to the STPK protein The importance of a dimer interface in phosphorylation (Fig. that the STPKs that are by phosphorylation with kinases with similar dimer Because of the of the interface residues of the three of M. tuberculosis receptor STPKs (3Gay L.M. Ng H.L. Alber T. J. Mol. Biol. 2006; 360: 409-420Crossref PubMed Scopus (51) Google it is that are formed between of the These support the that these three of STPKs that are by the of extracellular signaling and the of each signaling can bring together of the receptor kinases are to the roles of STPKs the of the signaling The allosteric activation mechanism of PknD from the of by dimerization of eukaryotic receptor kinases J. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). PknD activation with the allosteric regulation mechanisms of the human dsRNA-dependent (7Dar A.C. Dever T.E. Sicheri F. Cell. 2005; 122: 887-900Abstract Full Text Full Text PDF PubMed Scopus (287) Google Scholar, 8Dey M. Cao C. Dar A.C. Tamura T. Ozato K. Sicheri F. Dever T.E. Cell. 2005; 122: 901-913Abstract Full Text Full Text PDF PubMed Scopus (267) Google Scholar) and the EGF receptor kinase J. K. Kuriyan J. Cell. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). In response to binding of to the N-terminal ligand-binding domain, the KD a structurally similar back-to-back dimer (Fig. and Fig. which activates the kinase domains by phosphorylation of the activation (7Dar A.C. Dever T.E. Sicheri F. Cell. 2005; 122: 887-900Abstract Full Text Full Text PDF PubMed Scopus (287) Google Scholar, 8Dey M. Cao C. Dar A.C. Tamura T. Ozato K. Sicheri F. Dever T.E. Cell. 2005; 122: 901-913Abstract Full Text Full Text PDF PubMed Scopus (267) Google Scholar). Mutation of dimer interface from the active in a of activation and substrate phosphorylation M. Cao C. Dar A.C. Tamura T. Ozato K. Sicheri F. Dever T.E. Cell. 2005; 122: 901-913Abstract Full Text Full Text PDF PubMed Scopus (267) Google Scholar). in the dimer as and allosteric similar mechanism was by the dimer structure of which a distinct dimer through the N-terminal lobe A. J. Biol. Chem. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar). PknD the that the unphosphorylated dimer be active in a cognate substrate (Fig. and the phosphorylated kinase is active (Fig. through transmembrane the mechanism of activation for human epidermal growth receptor Fig. EGF binding to the extracellular receptor domain activates the intracellular KD by an dimer in which KD the of the other in the active J. K. Kuriyan J. Cell. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). This a distinct from the back-to-back dimer interface in (Fig. and PknD (Fig. In the kinase domain is to be between the PknD and activation a of are the of the allosteric interface is structurally conserved between and the M. tuberculosis receptor kinases (Fig. In the Src binding of of the linker domain to the analogous KD the inactive A. S. A. J. Mol. Biol. 1997; PubMed Scopus Google Scholar). These that this of the to the active site serves as an allosteric regulatory site in a of protein kinases. The of this interface in diverse STPKs implies that back-to-back dimerization a allosteric regulatory mechanism. The of this regulatory in kinases that the mechanisms of or from the allosteric to the active site are to be are to and for the of the FKBP and FRAP and Kuriyan provided