4.7 Article

NADP/H binding nearly doubles the stability of a Mycobacterium drug target: an unfolding study

Journal

JOURNAL OF BIOMOLECULAR STRUCTURE & DYNAMICS
Volume 41, Issue 16, Pages 8018-8025

Publisher

TAYLOR & FRANCIS INC
DOI: 10.1080/07391102.2022.2127910

Keywords

Mycobacterium tuberculosis; Aspartate beta semialdehyde dehydrogenase; NADP/H; tuberculosis; stability; unfolding; guanidinium chloride

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This study investigated the unfolding of Mycobacterium Aspartate beta semialdehyde dehydrogenase (ASADH) using spectroscopic techniques and size exclusion chromatography. The unfolding of the apo ASADH was found to be non-cooperative, while the unfolding of the holoenzyme was cooperative. The presence of NADP/H stabilizes the tryptophan environment and the native NADP/H-bound enzyme. The folded holoenzyme was shown to be conformationally more stable compared to the apo state. These findings are important for the discovery of novel drugs targeting ASADH.
Mycobacterium Aspartate beta semialdehyde dehydrogenase (ASADH) was studied using various spectroscopic techniques and size exclusion chromatography to examine the unfolding of free (apo) and NADP/H-bound (holo) forms of ASADH. Non-cooperative guanidinium chloride (GdnHCl)-induced unfolding of the apo ASADH was discovered, and no partially folded intermediate structures were stabilized. On the other hand, it was observed that GdnHCl's unfolding of holoenzyme was a cooperative process without any stable intermediate structure. The native form of holoenzyme is found to be stable against the lower concentration of GdnHCl only (namely up to 1.25M GdnHCl). The tryptophan environment appears to unfold cooperatively in case of the holoenzyme and is in well coordination with the overall unfolding of the holoenzyme. The presence of NADP/H shows a stabilizing effect on the tryptophan environment as well as on the native NADP/H-bound enzyme. Delta G(Solvent)(o) values reveal nearly two-fold (similar to 1.9) conformationally more stable folded holoenzyme compared to its native apo state. The C-m for the apo and holo forms of ASADH are 1.3 and 1.9M, respectively. Novel drug leads targeting the NADP/H binding domain of ASADH could offer promising drugs against extremely infective Mycobacterium tuberculosis.

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