2.8.1. Physiochemical Parameters

Mycobacterial virulent proteins are the major factors that aid in pathogenesis. The amino acid sequence of virulent proteins was obtained from the Mycobrowser database [55]. The Protparam tool was used to calculate the physicochemical parameters for these proteins, as shown in Table S4 [56]. The physiochemical parameter of the proteins calculated as the isoelectric points (pIs) of *Rv1477*, *Rv1495*, *Rv1566* and *Rv2801c* were greater than 7 (pI > 7), which means these proteins had more basic amino acids. The instability index value was also calculated and four unstable proteins (Instability index > 40) were found, viz. *Rv1477*, *Rv1566c*, *Rv2010*, and *Rv2623*. The calculated aliphatic index of some of these proteins were very high, which means they were thermostable. The GRAVY (grand average hydropathy) value was also determined, which indicated that *Rv0554*, *Rv1636*, *Rv2010*, and *Rv2623* were polar, and rest of the six proteins were non-polar (Figure 4a).

**Figure 4.** The selected virulence, detoxification, adaptation (VDA) category of *M. tuberculosis* H37Rv proteins. (**a**) Physiochemical parameters of VDA proteins (**b**) Functional classification of VDA proteins (**c**) Subcellular localization of VDA proteins and (**d**) Secondary structure analysis of VDA proteins.
