Structural and Biochemical Studies on Product Inhibition of S-Adenosylmethionine Synthetase from Corynebacterium glutamicum

Abstract

S-Adenosylmethionine (SAM) acts as a methyl donor in living organisms, and S-adenosylmethionine synthetase (MetK) is an essential enzyme for cells, as it synthesizes SAM from methionine and adenosine triphosphate (ATP). This study determined the crystal structures of the apo form and adenosine/triphosphate complex form of MetK from Corynebacterium glutamicum (CgMetK). Results showed that CgMetK has an allosteric inhibitor binding site for the SAM product in the vicinity of the active site and is inhibited by SAM both competitively and noncompetitively. Through structure-guided protein engineering, the CgMetK E68A variant was developed that exhibited an almost complete release of inhibition by SAM with rather enhanced enzyme activity. The crystal structure of the CgMetK E68A variant revealed that the formation of a new hydrogen bond between Tyr66 and Glu102 by the E68A mutation disrupted the allosteric SAM binding site and also improved the protein thermal stability by strengthening the tetramerization of the enzyme.

Engineering of Methionine Adenosyltransferase toward Mitigated Product Inhibition for Efficient Production of S-Adenosylmethionine

“…Generally, product inhibition is a pivotal factor that restricts the catalytic efficiency of MAT. The MAT from Corynebacterium glutamicum was engineered based on crystal structure analysis to mitigate the product inhibitory effects, and variant E68A was obtained with increased inhibition constant ( K i ) from 0.024 to 0.88 mM, leading to a significantly alleviated product inhibitory effect …”