The protein kinase PknB negatively regulates biosynthesis and trafficking of mycolic acids in mycobacteria

Abstract

Mycobacterium tuberculosis is the causative agent of tuberculosis and remains one of the most widespread and deadliest bacterial pathogens in the world. A distinguishing feature of mycobacteria that sets them apart from other bacteria is the unique architecture of their cell wall, characterized by various species-specific lipids, most notably mycolic acids (MAs). Therefore, targeted inhibition of enzymes involved in MA biosynthesis, transport, and assembly has been extensively explored in drug discovery. Additionally, more recent evidence suggests that many enzymes in the MA biosynthesis pathway are regulated by kinase-mediated phosphorylation, thus opening additional drug development opportunities. However, how phosphorylation regulates MA production remains unclear. Here, we employed genetic strategies combined with lipidomics and phosphoproteomics approaches to investigate the role of protein phosphorylation in Mycobacterium. The results of this analysis revealed that the Ser/Thr protein kinase PknB regulates export of MAs and promotes remodeling of the mycobacterial cell envelope. In particular, we identified the essential mycobacterial membrane protein large 3 (MmpL3) as a substrate negatively regulated by PknB. Taken together, our findings add to the understanding of how PknB activity affects the mycobacterial MA biosynthesis pathway and reveal the essential role of protein phosphorylation/dephosphorylation in governing lipid metabolism, paving the way towards novel antimycobacterial strategies.

Identification of residues important for M. tuberculosis MmpL11 function reveals that function is modulated by phosphorylation in the C‐terminal domain

“…Targets of PknB in M. smegmatis were identified through the overexpression of PknB (Le et al, 2020). Lipid analysis demonstrated that mycolic acid biosynthesis was impacted upon overexpression of PknB, with the accumulation of TMM and reduction of TDM levels.…”

Cg1246, a new player in mycolic acid biosynthesis in Corynebacterium glutamicum

“…In C. glutamicum, MA biosynthesis is regulated by transcriptional factors that either influence fatty acid synthesis (FasR [25,26], WhcD [27]), or control the expression of genes involved in MA metabolism (WhiA [28], σ D [29,30]). In mycobacteria a large number of enzymes involved in MA biosynthesis and transport were shown to be regulated by phosphorylation/dephosphorylation [31,32]. Although not demonstrated in C. glutamicum, it is very likely that this type of protein modification also contributes to fine-tuning some enzymatic and transport activities involved in corynomycolate metabolism in this bacterium.…”

Mycobacterium tuberculosis Thymidylyltransferase RmlA Is Negatively Regulated by Ser/Thr Protein Kinase PknB

“…PknB, a well-known M. tuberculosis STPK, is believed to regulate cell wall biosynthesis and cell division ( Banu et al, 2010 ). The extracellular domain of PknB binds peptidoglycan-derived muropeptides for its localization ( Roumestand et al, 2011 ), and several phosphorylated substrates have been identified, such as SahH ( Singhal et al, 2013 ), InhA ( Vilcheze and Jacobs, 2014 ; Shaw et al, 2017 ), KasB ( Vilcheze et al, 2014 ), GlmU ( Parikh et al, 2009 ), MmpL3 ( Le et al, 2020 ), CwIM ( Turapov et al, 2018 ), and Ef-Tu ( Sajid et al, 2011 ), in participating in cell metabolic, cell wall synthesis, transcription, and translation processes.…”