Reversible, partial inactivation of plant betaine aldehyde dehydrogenase by betaine aldehyde: mechanism and possible physiological implications

Abstract

In plants, the last step in the biosynthesis of the osmoprotectant glycine betaine (GB) is the NAD+-dependent oxidation of betaine aldehyde (BAL) catalysed by some aldehyde dehydrogenase (ALDH) 10 enzymes that exhibit betaine aldehyde dehydrogenase (BADH) activity. Given the irreversibility of the reaction, the short-term regulation of these enzymes is of great physiological relevance to avoid adverse decreases in the NAD+:NADH ratio. In the present study, we report that the Spinacia oleracea BADH (SoBADH) is reversibly and partially inactivated by BAL in the absence of NAD+ in a time- and concentration-dependent mode. Crystallographic evidence indicates that the non-essential Cys450 (SoBADH numbering) forms a thiohemiacetal with BAL, totally blocking the productive binding of the aldehyde. It is of interest that, in contrast to Cys450, the catalytic cysteine (Cys291) did not react with BAL in the absence of NAD+. The trimethylammonium group of BAL binds in the same position in the inactivating or productive modes. Accordingly, BAL does not inactivate the C450S SoBADH mutant and the degree of inactivation of the A441I and A441C mutants corresponds to their very different abilities to bind the trimethylammonium group. Cys450 and the neighbouring residues that participate in stabilizing the thiohemiacetal are strictly conserved in plant ALDH10 enzymes with proven or predicted BADH activity, suggesting that inactivation by BAL is their common feature. Under osmotic stress conditions, this novel partial and reversible covalent regulatory mechanism may contribute to preventing NAD+ exhaustion, while still permitting the synthesis of high amounts of GB and avoiding the accumulation of the toxic BAL.