Mycobacterium fortuitum fabG4knockdown studies: Implication as pellicle and biofilm specific drug target

Abstract

The fatty acid biosynthesis pathway is crucial for the formation of the mycobacterial cell envelope. The fatty acid synthase type-II (FAS-II) components are attractive targets for designing anti-biofilm inhibitors.Literature review, bioinformatics analysis, cloning, and sequencing led to the identification of a novel Mycobacterium fortuitum FAS-II gene MFfabG4 which interacts with mycobacterial proteins involved in biofilm formation. A manually curated M. fortuitum fatty acid biosynthesis pathway has been proposed exploiting functional studies from the Kyoto Encyclopedia of Genes and Genomes and Mycobrowser databases for MFFabG4. M. fortuitum MFfabG4 knockdown strain (FA) was constructed and validated by quantitative polymerase chain reaction. The FA strain displayed unstructured smooth colony architecture, correlating with decreased pathogenicity and virulence. MFfabG4 knockdown resulted in diminished pellicle and attenuated biofilm formation, along with impaired sliding motility, and reduced cell sedimentation. The FA strain showed lowered cell surface hydrophobicity, indicating attenuation in M. fortuitum intracellular infection-causing ability. Stress survival studies showed the requirement of MFfabG4 for survival in a nutrient-starved environment. The results indicate that MFfabG4 maintains the physiology of the cell envelope and is required for the formation of M. fortuitum pellicle and biofilm. The study corroborates the role of MFfabG4 as a pellicle-and biofilm-specific drug target and a potential diagnostic marker for M. fortuitum and related pathogenic mycobacteria.

Mycobacterium biofilms

“…2 ) [ 18 , 19 ]. Other molecules involved in biofilm development have been described, such as glycopeptidolipids (GPLs) [ [20] , [21] , [22] , [23] , [24] ], type III polyketide synthases [ 25 ], ESX-1 secretion system [ 26 ], GroEL1 chaperone [ 27 ], FabG4 [ 28 ], Peptidyl-prolyl isomerase [ 29 ], FAS-II components [ 30 ], protein kinase PknF [ 31 ] and many others in different mycobacterial species, most of them rapidly growing [ [32] , [33] , [34] ]. Most of these molecules and systems are involved in the mycolic acid synthesis and the metabolism of different components of the mycobacterial cell wall.…”

Structural and functional characterization of FabG4 from Mycolicibacterium smegmatis

“…In M. tuberculosis, FabG4 was found to be expressed under limited nutrient conditions and in the presence of antibiotic stressors, suggesting its involvement in drug resistance in mycobacterial species (Beste et al, 2009;Sharma et al, 2010). FabG4 is also found in the cell-envelope membrane fraction of M. tuberculosis and is hypothesized to be involved in mycobacterial biofilm formation (Sharma et al, 2019(Sharma et al, , 2022. In A. baumannii, HMwFabG was found to be essential in several processes, including motility, pellicle formation and growth in nutrient-rich broth/conditions (Cross et al, 2021).…”