丝氨酸/苏氨酸蛋白激酶在结核分枝杆菌致病机制中的作用

康涵，吴文娟

上海市(复旦大学附属)公共卫生临床中心，上海 201508

丝氨酸/苏氨酸蛋白激酶(serine/threonine protein kinase，STPK)是一类真核细胞样的蛋白激酶，是分枝杆菌生长和代谢的重要调节因子。结核分枝杆菌共有11种STPK，分为跨膜蛋白和可溶性蛋白两大类[1]。对多种分枝杆菌STPK基因序列进行分析，按系统发生可分为5群：ABL 群(pknA、pknB、pknL)，HED群(pknH、pknE、pknD)，FIJ群(pknF、pknI、pknJ)，pknG及pknK[2]。研究发现，STPK参与分枝杆菌多种细胞活动(如细胞和菌落形态、葡萄糖和谷氨酰胺转运、吞噬体-溶酶体融合、转录因子活性等)的调节(表1)。STPK与其配体结合，二聚化后构象改变引起自身磷酸化，并磷酸化下游蛋白，从而发挥信号转导作用[3]。Cui等[4]对结核分枝杆菌蛋白间相互作用的预测显示，STPK可与许多重要蛋白发生相互作用。Malhotra等[5]对结核分枝杆菌H37Rv和H37Ra的STPK基因表达水平比较分析发现，有毒株H37Rv的6个STPK基因pknD、pknG、pknH、pknJ、pknK和pknL有更高的表达。

表1 结核分枝杆菌的丝氨酸/苏氨酸蛋白激酶
Fig.1 The serine/threonine protein kinases ofMycobacteriumtuberculosis

NameORFLengthClassificationRegulatory role PknARv0015c431 aaTransmembrane Cell division and morphologyPknBRv0014c626 aaTransmembraneCell division and morphologyPknLRv2176399 aaTransmembranePhosphorylation of Rv2175c, transcription?PknHRv1266c626 aaTransmembraneArabinan metabolism, growth in hostile environmentPknERv1743566 aaTransmembraneResponse to NO pressure, apoptosis of infected macrophagesPknDRv0931c664 aaTransmembraneTranscription, phosphate transport?PknFRv1746476 aaTransmembraneCell divisionPknIRv2914c585 aaTransmembraneGrowth in hostile environmentPknJRv2088589 aaTransmembranePhosphorylation of sorts of functional proteinsPknGRv0410c750 aaSolubleGlutamate/glutamine metabolism, intrinsic antibiotic resistance, in-hibition of phagosome-lysosome fusion, enhancing bacterial intra-cellular survivalPknKRv3080c1110 aaSolubleTranscription, cell wall ultrastructure, growth in vivo

1 跨膜STPK

1.1 ABL群

目前研究表明，PknA和PknB主要与细胞形态和分裂有关，PknL可磷酸化DNA结合蛋白Rv2175c。Schultz等[6]构建了多种谷氨酸棒状杆菌的STPK突变株，发现PknA、PknB、PknG和PknL对细菌的生存不是必需的，它们都能磷酸化2-酮戊二酸脱氢酶OdhI，其中又以PknG作用最强；此外，细胞分裂蛋白FtsZ也能被PknA、PknB和PknL磷酸化。但Fiuza等[7]认为，PknA和 PknB对谷氨酸棒状杆菌的生长是必需的，其过表达使杆菌呈小球状生长。在结核分枝杆菌中，PknA和PknB也有相似作用：PknA能磷酸化细胞分裂蛋白FtsZ及分枝杆菌UDP-N-乙酰胞壁酰-L-丙氨酰-D-谷氨酸连接酶[8, 9]，PknB也有多种底物，包括GarA、N-乙酰葡糖胺-1-磷酸尿苷酰转移酶、叉头结构域相关蛋白Rv0019c、PapA5等[10-12]；而PknA和PknB能被跨膜的Mstp蛋白去磷酸化[13]。这些底物多与分枝杆菌细胞壁合成、细胞分裂等有关，在大肠埃希菌中表达结核分枝杆菌pknA可引起细菌异常延伸[14]，表达结核分枝杆菌pknA和pknB的耻垢分枝杆菌生长变慢，敲除pknA和pknB的结核分枝杆菌菌体变得更狭窄[15]。PknL可磷酸化一种放线菌的保守DNA结合蛋白Rv2175c[16,17]，但其下游效应还不清楚。

1.2 HED群

此群STPK参与细菌的多种重要生理活动，并与致病性密切相关。PknH存在于致病的结核分枝杆菌和牛分枝杆菌，而不存在于耻垢分枝杆菌中。在压力环境下，结核分枝杆菌pknH的表达水平改变[18]。重组表达pknH的耻垢分枝杆菌中，PknH能磷酸化EmbR，导致embCAB操纵子的转录水平增加，影响阿拉伯糖代谢，使脂化甘露聚醣与脂化阿拉伯甘露聚糖的比值升高，并对乙胺丁醇的抵抗性增加。结核分枝杆菌感染巨噬细胞后，pknH表达增高[19]。但另有研究发现，在结核分枝杆菌中敲除pknH并不影响embCAB操纵子的表达，仅减少乙胺丁醇诱导的embCAB操纵子的表达[20]。除EmbR外，Zheng等[21]在体外实验中发现，PknH还能磷酸化Rv0681和DacB1。敲除pknH的结核分枝杆菌感染BALB/c小鼠后，在感染慢性期生存和复制能力增强[20]。对PknE的活性、定位、晶体结构及结构域的分析提示，它可能是跨膜的受体型蛋白激酶，能接收细胞外的信号并将其传递到细胞内，引起细胞活动改变。该过程主要与其结合配体后二聚化变构并自身磷酸化有关[22, 23]。体外实验发现，PknE、PknD均能磷酸化ATP结合盒转运蛋白(ATP-binding cassette transporter, ABC)[24]。PknE在结核分枝杆菌对一氧化氮(nitric oxide，NO)压力反应及人巨噬细胞感染后的凋亡有重要作用。与野生株相比，pknE突变株对NO供体的抵抗力增强，而对还原剂和某些金属离子的抵抗力减弱，引起巨噬细胞凋亡增加，但巨噬细胞坏死和增殖减少[25]。结核分枝杆菌的PknD跨膜区C端有个β-propeller模体，而牛分枝杆菌中无此结构。结核分枝杆菌毒力相关转运蛋白MmpL7可能是PknD的底物之一[26]。PknD也许与磷酸盐转运有关，但尚未证实[27,28]。PknD具有与PknE相似的二聚化及变构激活过程[29]。体外实验发现，PknD过表达能改变多种基因转录，包括Rv0516c及一些受sigma因子F调节的基因等[30]。

1.3 FIJ群

目前研究显示，PknF与细菌分裂有关，PknI参与调节有害环境中的细菌生长，PknJ则可磷酸化多种功能蛋白，进而影响细菌的代谢活动。体外实验发现，PknF也可磷酸化多种含有叉头相关结构域的蛋白，如ABC转运子、髓磷脂碱性蛋白等[24,31,32]。PknF在结核分枝杆菌葡萄糖转化、细胞生长及隔膜形成中可能起直接或间接作用[33]。PknF也表达于耻垢分枝杆菌，它能影响耻垢分枝杆菌的生物膜形成及运动性，这与结核分枝杆菌相似[34]。PknI虽然为跨膜STPK，但有报道发现其也存在于细胞质中[35]。基因信息分析预测，PknI可能参与细胞生长和延伸[36]。感染巨噬细胞后，pknI表达下调[33]。与野生株相比，PknI突变株在酸性条件及缺氧时生长更好，感染巨噬细胞后其生存力更强，在免疫缺陷小鼠中突变株的毒力也更高[37]。PknJ的底物较多，包括PknJ本身、转录调节因子EmbR、参与分枝菌酸合成的甲基转移酶MmaA4/Hma、二肽酶PepE、丙酮酸激酶A等[38, 39]。

2 可溶性STPK

2.1 PknG

PknG与真核细胞的蛋白激酶Cα(protein kinase Cα, PKCα)很相似，可自我磷酸化，其磷酸化能被白屈菜赤碱阻断。与其他大多数结核分枝杆菌的STPK不同，PknG中未发现已知的穿膜结构域。耻垢分枝杆菌不表达PknG，结核分枝杆菌和牛分枝杆菌卡介苗(bacillus Calmette-Guerin，BCG)表达PknG[40]。亚细胞定位发现，PknG存在于分枝杆菌的胞质溶胶和细胞膜中[41]，巨噬细胞摄入分枝杆菌后，在吞噬体和胞质溶胶内都能检测到PknG存在[40]。结核分枝杆菌pknG缺陷株可造成谷氨酸盐和谷氨酰胺堆积，使其细胞内浓度达到野生株的3倍[41]。但Nguyen等发现，BCG中pknG缺陷并不影响谷氨酰胺的摄入及其细胞内浓度，他们认为BCG中的谷氨酰胺代谢不受PknG调节[42]。谷氨酸盐和谷氨酰胺在分枝杆菌中的作用包括：①谷氨酸盐和谷氨酰胺的合成及潜在的脱氢酶活性是分枝杆菌氨基酸氨基同化作用的唯一途径；②聚合L-谷氨酸盐/谷氨酰胺是结核分枝杆菌细胞膜结构完整性的一部分[41]。PknG缺陷造成谷氨酸盐/谷氨酰胺过度堆积可能是影响分枝杆菌生存和毒力的机制之一。比较结核分枝杆菌pknG缺陷株与野生株对多种抗生素的最低抑菌浓度发现，PknG在结核分枝杆菌的天然耐药中起重要作用[43]。体外生长实验发现，PknG对BCG体外生存不是必需的，突变株和野生株在细胞形态和生长上都没有差别，其特异性抑制剂对分枝杆菌在培养物中的生长亦没有影响[40,42]。而体内或巨噬细胞受结核分枝杆菌感染后，情况则有所不同，重症联合免疫缺陷(severe combined immunodeficiency，SCID)小鼠感染突变株后死亡时间推迟，且突变株在SCID及BALB/c小鼠体内的生存力均下降。这在分枝杆菌生长周期的静止相或营养缺乏时尤为显著[41]。导入pknG可延长耻垢分枝杆菌在巨噬细胞内的存活。BCG感染巨噬细胞后，突变株主要存在于吞噬溶酶体内，而野生株则主要存在于非溶酶体细胞器内。PknG特异抑制剂可导致剂量依赖的、分枝杆菌感染的巨噬细胞内吞噬溶酶体的形成，吞噬体迅速与溶酶体融合并杀死其中的分枝杆菌。进一步研究发现，这些与PknG的激酶活性有关[40]。此外，H37Rv、H37Ra及BCG感染巨噬细胞后，都发现PknG能下调巨噬细胞PKCα表达，从而影响巨噬细胞对分枝杆菌的吞噬，并增加分枝杆菌在细胞内存活。重组了pknG的耻垢分枝杆菌在巨噬细胞内的生存能力也显著增强[44]。

2.2 PknK

与PknG不同，PknK主要存在于细胞壁和细胞膜(后者杂交信号很弱)，在感染巨噬细胞中仅溶酶体内有信号检出[5, 45]。已知PknK底物包括VirS和mymA操纵子编码的4个蛋白。PknK能磷酸化转录因子VirS，增强其与DNA的结合能力，进而引起mymA的转录及表达[45]。mymA操纵子编码多个酶，参与分枝杆菌脂类代谢。virS和(或)mymA敲除的结核分枝杆菌细胞壁超微结构发生改变，分枝菌酸的合成受影响；对去污剂、酸性环境及抗生素的敏感性增加；在活化巨噬细胞中的生存能力及对豚鼠的毒力也发生改变[46]。扫描电镜观察发现，pknK突变株比野生株菌体小，细胞壁薄。仅在结核分枝杆菌H37Rv感染巨噬细胞后18 h，pknK有显著表达。体外生长实验发现，PknK能抑制结核分枝杆菌在稳定期的生长。感染C57BL/6小鼠后，突变株在感染早期生长受限而后期增强；但在静息巨噬细胞内，突变株与野生株的生长情况相似，说明其在小鼠体内生长不同并不是由于其内在的感染性不同。分析不同时间点细胞因子的mRNA 表达发现，在突变株感染早期，白细胞介素2(interleukin 2，IL-12)、肿瘤坏死因子α(tumor necrosis factor α，TNF-α)、γ干扰素(interferon γ，IFN-γ)、诱导型一氧化氮合酶(inducible nitric oxide synthase，iNOS)的表达均低于野生株，后期则差异不显著。表明PknK可能在调节体内环境压力下的生长中起重要作用[5]。

3 结语

面对结核病对人类健康的重大挑战，结核分枝杆菌致病机制的明确将给抗结核病带来重大突破，如新药物靶点的发现、新型疫苗的开发等。虽然已有大量研究发现了结核病相关致病物质[47-49]，但具体机制尚不明确。STPK在结核分枝杆菌的许多细胞活动中起重要作用，能感知细菌所处的环境并作出反应，且可能与结核分枝杆菌-宿主免疫平衡的维持有关。对于多数STPK，其作用机制还很不清楚，需进一步研究。

[1] Cole ST, Brosch R, Parkhill J, Garnier T, Churcher C, Harris D, Gordon SV, Eiglmeier K, Gas S, Barry CE 3rd, Tekaia F, Badcock K, Basham D, Brown D, Chillingworth T, Connor R, Davies R, Devlin K, Feltwell T, Gentles S, Hamlin N, Holroyd S, Hornsby T, Jagels K, Krogh A, McLean J, Moule S, Murphy L, Oliver K, Osborne J, Quail MA, Rajandream MA, Rogers J, Rutter S, Seeger K, Skelton J, Squares R, Squares S, Sulston JE, Taylor K, Whitehead S, Barrell BG. Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence [J]. Nature, 1998, 393(6685): 537-544.

[2] Narayan A, Sachdeva P, Sharma K, Saini AK, Tyagi AK, Singh Y. Serine threonine protein kinases of mycobacterial genus: phylogeny to function [J]. Physiol Genomics, 2007, 29(1): 66-75.

[3] Alber T. Signaling mechanisms of the Mycobacterium tuberculosis receptor Ser/Thr protein kinases [J]. Curr Opin Struct Biol, 2009, 19(6): 650-657.

[4] Cui T, Zhang L, Wang X, He ZG. Uncovering new signaling proteins and potential drug targets through the interactome analysis of Mycobacterium tuberculosis [J]. BMC Genomics, 2009, 10: 118.

[5] Malhotra V, Arteaga-Cortés LT, Clay G, Clark-Curtiss JE. Mycobacterium tuberculosis protein kinase K confers survival advantage during early infection in mice and regulates growth in culture and during persistent infection: implications for immune modulation [J]. Microbiology, 2010, 156(Pt 9): 2829-2841.

[6] Schultz C, Niebisch A, Schwaiger A, Viets U, Metzger S, Bramkamp M, Bott M. Genetic and biochemical analysis of the serine/threonine protein kinases PknA, PknB, PknG and PknL of Corynebacterium glutamicum: evidence for non-essentiality and for phosphorylation of OdhI and FtsZ by multiple kinases [J]. Mol Microbiol, 2009, 74(3): 724-741.

[7] Fiuza M, Canova MJ, Zanella-Cléon I, Becchi M, Cozzone AJ, Mateos LM, Kremer L, Gil JA, Molle V. From the characterization of the four serine/threonine protein kinases (PknA/B/G/L) of Corynebacterium glutamicum toward the role of PknA and PknB in cell division [J]. J Biol Chem, 2008, 283(26): 18099-18112.

[8] Thakur M，Chakraborti PK. GTPase activity of mycobacterial FtsZ is impaired due to its transphosphorylation by the eukaryotic-type Ser/Thr kinase, PknA [J]. J Biol Chem, 2006, 281(52): 40107-40113.

[9] Thakur M，Chakraborti PK. Ability of PknA, a mycobacterial eukaryotic-type serine/threonine kinase, to transphosphorylate MurD, a ligase involved in the process of peptidoglycan biosynthesis [J]. Biochem J, 2008, 415(1): 27-33.

[10] Gupta M, Sajid A, Arora G, Tandon V, Singh Y. Forkhead-associated domain-containing protein Rv0019c and polyketide-associated protein PapA5, from substrates of serine/threonine protein kinase PknB to interacting proteins of Mycobacterium tuberculosis [J]. J Biol Chem, 2009, 284(50): 34723-34734.

[11] Parikh A, Verma SK, Khan S, Prakash B, Nandicoori VK. PknB-mediated phosphorylation of a novel substrate, N-acetylglucosamine-1-phosphate uridyltransferase, modulates its acetyltransferase activity [J]. J Mol Biol, 2009, 386(2): 451-464.

[12] Villarino A, Duran R, Wehenkel A, Fernandez P, England P, Brodin P, Cole S T,Zimny-Arndt U, Jungblut PR, Cervenansky C, Alzari PM. Proteomic identification of M. tuberculosis protein kinase substrates: PknB recruits GarA, a FHA domain-containing protein, through activation loop-mediated interactions [J]. J Mol Biol, 2005, 350(5): 953-963.

[13] Chopra P, Singh B, Singh R, Vohra R, Koul A, Meena LS, Koduri H, Ghildiyal M, Deol P, Das TK, Tyagi AK, Singh Y. Phosphoprotein phosphatase of Mycobacterium tuberculosis dephosphorylates serine-threonine kinases PknA and PknB [J]. Biochem Biophys Res Commun, 2003, 311(1): 112-120.

[14] Chaba R, Raje M, Chakraborti PK. Evidence that a eukaryotic-type serine/threonine protein kinase from Mycobacterium tuberculosis regulates morphological changes associated with cell division [J]. Eur J Biochem, 2002, 269(4): 1078-1085.

[15] Kang CM, Abbott DW, Park ST, Dascher CC, Cantley LC, Husson RN. The Mycobacterium tuberculosis serine/threonine kinases PknA and PknB: substrate identification and regulation of cell shape[J]. Genes Dev, 2005, 19(14): 1692-1704.

[16] Canova MJ, Veyron-Churlet R, Zanella-Cleon I, Cohen-Gonsaud M, Cozzone AJ, Becchi M, Kremer L, Molle V. The Mycobacterium tuberculosis serine/threonine kinase PknL phosphorylates Rv2175c: mass spectrometric profiling of the activation loop phosphorylation sites and their role in the recruitment of Rv2175c [J]. Proteomics, 2008, 8(3): 521-533.

[17] Cohen-Gonsaud M, Barthe P, Canova MJ, Stagier-Simon C, Kremer L, Roumestand C, Molle V. The Mycobacterium tuberculosis Ser/Thr kinase substrate Rv2175c is a DNA-binding protein regulated by phosphorylation [J]. J Biol Chem, 2009, 284(29): 19290-19300

[18] Sharma K, Chandra H, Gupta PK, Pathak M, Narayan A, Meena LS, D’Souza RC, Chopra P, Ramachandran S, Singh Y. PknH, a transmembrane Hank’s type serine/threonine kinase from Mycobacterium tuberculosis is differentially expressed under stress conditions [J]. FEMS Microbiol Lett, 2004, 233(1): 107-113.

[19] Sharma K, Gupta M, Pathak M, Gupta N, Koul A, Sarangi S, Baweja R, Singh Y. Transcriptional control of the mycobacterial embCAB operon by PknH through a regulatory protein, EmbR, in vivo [J]. J Bacteriol, 2006, 188(8): 2936-2944.

[20] Papavinasasundaram KG, Chan B, Chung JH, Colston MJ, Davis EO, Av-Gay Y. Deletion of the Mycobacterium tuberculosis pknH gene confers a higher bacillary load during the chronic phase of infection in BALB/c mice [J]. J Bacteriol, 2005, 187(16): 5751-5760.

[21] Zheng X, Papavinasasundaram KG, Av-Gay Y. Novel substrates of Mycobacterium tuberculosis PknH Ser/Thr kinase [J]. Biochem Biophys Res Commun, 2007, 355(1): 162-168.

[22] Molle V, Girard-Blanc C, Kremer L, Doublet P, Cozzone AJ, Prost JF. Protein PknE, a novel transmembrane eukaryotic-like serine/threonine kinase from Mycobacterium tuberculosis [J]. Biochem Biophys Res Commun, 2003, 308(4): 820-825.

[23] Gay LM, Ng HL, Alber T. A conserved dimer and global conformational changes in the structure of apo-PknE Ser/Thr protein kinase from Mycobacterium tuberculosis [J]. J Mol Biol, 2006, 360(2): 409-420.

[24] Grundner C, Gay LM, Alber T. Mycobacterium tuberculosis serine/threonine kinases PknB, PknD, PknE, and PknF phosphorylate multiple FHA domains [J]. Protein Sci, 2005, 14(7): 1918-1921.

[25] Jayakumar D, Jacobs WR Jr, Narayanan S. Protein kinase E of Mycobacterium tuberculosis has a role in the nitric oxide stress response and apoptosis in a human macrophage model of infection [J]. Cell Microbiol, 2008, 10(2): 365-374.

[26] Pérez J, Garcia R, Bach H, de Waard JH, Jacobs WR Jr, Av-Gay Y, Bubis J, Takiff HE. Mycobacterium tuberculosis transporter MmpL7 is a potential substrate for kinase PknD [J]. Biochem Biophys Res Commun, 2006, 348(1): 6-12.

[27] Av-Gay Y, Everett M. The eukaryotic-like Ser/Thr protein kinases of Mycobacterium tuberculosis [J]. Trends Microbiol, 2000, 8(5): 238-244.

[28] Lefèvre P, Braibant M, de Wit L, Kalai M, Röeper D, Grötzinger J, Delville JP, Peirs P, Ooms J, Huygen K, Content J. Three different putative phosphate transport receptors are encoded by the Mycobacterium tuberculosis genome and are present at the surface of Mycobacterium bovis BCG [J]. J Bacteriol, 1997, 179(9): 2900-2906.

[29] Greenstein AE, Echols N, Lombana TN, King DS, Alber T. Allosteric activation by dimerization of the PknD receptor Ser/Thr protein kinase from Mycobacterium tuberculosis [J]. J Biol Chem, 2007, 282(15): 11427-11435.

[30] Greenstein AE, MacGurn JA, Baer CE, Falick AM, Cox JS, Alber T. M. tuberculosis Ser/Thr protein kinase D phosphorylates an anti-anti-sigma factor homolog [J]. PLoS Pathog, 2007, 3(4): e49.

[31] Koul A, Choidas A, Tyagi AK, Drlica K, Singh Y, Ullrich A. Serine/threonine protein kinases PknF and PknG of Mycobacterium tuberculosis: characterization and localization [J]. Microbiology, 2001, 147(Pt 8): 2307-2314.

[32] Molle V, Soulat D, Jault JM, Grangeasse C, Cozzone AJ, Prost JF. Two FHA domains on an ABC transporter, Rv1747, mediate its phosphorylation by PknF, a Ser/Thr protein kinase from Mycobacterium tuberculosis [J]. FEMS Microbiol Lett, 2004, 234(2): 215-223.

[33] Deol P, Vohra R, Saini AK, Singh A, Chandra H, Chopra P, Das TK, Tyagi AK, Singh Y. Role of Mycobacterium tuberculosis Ser/Thr kinase PknF: implications in glucose transport and cell division [J]. J Bacteriol, 2005, 187(10): 3415-3420.

[34] Gopalaswamy R, Narayanan S, Jacobs WR Jr, Av-Gay Y. Mycobacterium smegmatis biofilm formation and sliding motility are affected by the serine/threonine protein kinase PknF [J]. FEMS Microbiol Lett, 2008, 278(1): 121-127.

[35] Singh A, Singh Y, Pine R, Shi L, Chandra R, Drlica K. Protein kinase I of Mycobacterium tuberculosis: cellular localization and expression during infection of macrophage-like cells [J]. Tuberculosis (Edinb), 2006, 86(1): 28-33.

[36] Gopalaswamy R, Narayanan PR, Narayanan S. Cloning, overexpression, and characterization of a serine/threonine protein kinase pknI from Mycobacterium tuberculosis H37Rv [J]. Protein Expr Purif, 2004, 36(1): 82-89.

[37] Gopalaswamy R, Narayanan S, Chen B, Jacobs WR, Av-Gay Y. The serine/threonine protein kinase PknI controls the growth of Mycobacterium tuberculosis upon infection [J]. FEMS Microbiol Lett, 2009, 295(1): 23-29.

[38] Arora G, Sajid A, Gupta M, Bhaduri A, Kumar P, Basu-Modak S, Singh Y. Understanding the role of PknJ in Mycobacterium tuberculosis: biochemical characterization and identification of novel substrate pyruvate kinase A [J]. PLoS One, 2010, 5(5): e10772.

[39] Jang J, Stella A, Boudou F, Levillain F, Darthuy E, Vaubourgeix J, Wang C, Bardou F, Puzo G, Gilleron M, Burlet-Schiltz O, Monsarrat B, Brodin P, Gicquel B, Neyrolles O. Functional characterization of the Mycobacterium tuberculosis serine/threonine kinase PknJ [J]. Microbiology, 2010, 156(Pt 6): 1619-1631.

[40] Walburger A, Koul A, Ferrari G, Nguyen L, Prescianotto-Baschong C, Huygen K, Klebl B, Thompson C, Bacher G, Pieters J. Protein kinase G from pathogenic mycobacteria promotes survival within macrophages [J]. Science, 2004, 304(5678): 1800-1804.

[41] Cowley S, Ko M, Pick N, Chow R, Downing KJ, Gordhan BG, Betts JC, Mizrahi V, Smith DA, Stokes RW, Av-Gay Y. The Mycobacterium tuberculosis protein serine/threonine kinase PknG is linked to cellular glutamate/glutamine levels and is important for growth in vivo [J]. Mol Microbiol, 2004, 52(6): 1691-1702.

[42] Nguyen L, Walburger A, Houben E, Koul A, Muller S, Morbitzer M, Klebl B, Ferrari G, Pieters J. Role of protein kinase G in growth and glutamine metabolism of Mycobacterium bovis BCG [J]. J Bacteriol, 2005, 187(16): 5852-5856.

[43] Wolff KA, Nguyen HT, Cartabuke RH, Singh A, Ogwang S, Nguyen L. Protein kinase G is required for intrinsic antibiotic resistance in mycobacteria [J]. Antimicrob Agents Chemother, 2009, 53(8): 3515-3519.

[44] Chaurasiya SK, Srivastava KK. Downregulation of protein kinase C-alpha enhances intracellular survival of Mycobacteria: role of PknG [J]. BMC Microbiol, 2009, 9: 271.

[45] Kumar P, Kumar D, Parikh A, Rananaware D, Gupta M, Singh Y, Nandicoori VK. The Mycobacterium tuberculosis protein kinase K modulates activation of transcription from the promoter of mycobacterial monooxygenase operon through phosphorylation of the transcriptional regulator VirS [J]. J Biol Chem, 2009, 284(17): 11090-11099.

[46] Singh A, Gupta R, Vishwakarma RA, Narayanan PR, Paramasivan CN, Ramanathan VD, Tyagi AK. Requirement of the mymA operon for appropriate cell wall ultrastructure and persistence of Mycobacterium tuberculosis in the spleens of guinea pigs [J]. J Bacteriol, 2005, 187(12): 4173-4186.

[47] Meena LS, Rajni. Survival mechanisms of pathogenic Mycobacterium tuberculosis H37Rv [J]. FEBS J, 2010, 277(11): 2416-2427.

[48] Tian C, Jian-Ping X. Roles of PE_PGRS family in Mycobacterium tuberculosis pathogenesis and novel measures against tuberculosis [J]. Microb Pathog, 2010, 49(6): 311-314.

[49] Ehrt S, Schnappinger D. Mycobacterial survival strategies in the phagosome: defence against host stresses [J]. Cell Microbiol, 2009, 11(8): 1170-1178.