Serine/ threonine protein kinases (STPKs) are known to act as sensors of environmental signals which thereby regulate developmental changes and host pathogen interactions. In this study, we carried out comparative genome analysis of six completely sequenced pathogenic and non-pathogenic mycobacterial species to systematically characterize the STPK complement of mycobacterium. Our analysis revealed that while M. tuberculosis strains have eleven conserved kinases, this number varies from four to twenty four in other mycobacterial species. pknA, an essential STPK encoding gene was found to be truncated in the initial analysis of M. avium subsp. paratuberculosis (Map) and M. tuberculosis C genomes. However, resequencing of pknA gene in Map confirmed that the truncation was due to a sequencing error. The conservation of division and cell wall (dcw) gene cluster, involved in cell envelope biosynthesis and cell division, in the vicinity of pknL locus, implicates a possible role of PknL in cell division and envelop biosynthesis. We identified a cyclophilin domain as part of a mycobacterial kinase in Map that suggests a plausible regulation of cyclophilins by phosphorylation. The co-inheritance of pknA, pknB, pknG and pknL loci across genomes and some unique repertoire of pathogen specific kinases such as pknI and pknJ of Mtb complex indicate towards the similitude and divergence between pathogenic and non pathogenic signaling. This study would add another dimension towards identification of physiological substrates and thereby function, while resolving the existing complexities in signaling network between the two domains of life, pathogen and non pathogen.