Functional importance of ?Ile-346 and ?Ile-348 in the catalytic sites of Escherichia coli ATP synthase.

ABSTRACT: We studied the functional role of highly conserved VISIT-DG sequence residues ?Ile-346 and ?Ile-348 in the catalytic sites of Escherichia coli F1Fo ATP synthase. ?Ile-346 is in close proximity, 2.98 and 3.63 Å, to the two known phosphate binding residues ?R376 and ?R182; ?Ile-348 is situated within 3.66 Å from ?R182. Single or double mutants of both ?I346 and ?I348 resulted in a variable loss of oxidative phosphorylation and ATPase activity. Azide, fluoroaluminate, and fluoroscandium caused insignificant to significant inhibition of mutants. Whereas the wild-type enzyme was completely inhibited by NBD-Cl (7-chloro-4-nitrobenzo-2-oxa-1, 3-diazole), a variable extent of inhibition was observed for ?I346 and ?I348 mutants. MgPi protection against NBD-Cl induced inhibition of wild-type, ?I346, and ?I348 demonstrated that, although strongly conserved, ?I346 and ?I348 have no direct role in phosphate binding. Insertion of Arginine in the form of ?I346R/?R182A, ?I346R/?R376A, or ?I348R/?R182A was able to compensate for the absence of known phosphate-binding Arginine residues ?R182 and ?R376. Results also suggest that ?Ile-346 and ?Ile-348 seem to have functional importance in upholding the phosphate-binding subdomain and transition state stabilization in the catalytic sites of E. coli ATP synthase.

SUBMITTER: Zhao C 

PROVIDER: S-EPMC5884630 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature