Promiscuous enzymes generating d-amino acids in mammals: Why they may still surprise us?

Promiscuous catalysis is a common property of enzymes, particularly those using pyridoxal 5′-phosphate as a cofactor. In a recent issue of this journal, Katane et al. Biochem. J. 477, 4221–4241 demonstrate the synthesis and accumulation of d-glutamate in mammalian cells by promiscuous catalysis mediated by a pyridoxal 5′-phosphate enzyme, the serine/threonine dehydratase-like (SDHL). The mechanism of SDHL resembles that of serine racemase, which synthesizes d-serine, a well-established signaling molecule in the mammalian brain. d-Glutamate is present in body fluids and is degraded by the d-glutamate cyclase at the mitochondria. This study demonstrates a biochemical pathway for d-glutamate synthesis in mammalian cells and advances our knowledge on this little-studied d-amino acid in mammals. d-Amino acids may still surprise us by their unique roles in biochemistry, intercellular signaling, and as potential biomarkers of disease.

A novel deep-sea bacterial threonine dehydratase drives cysteine desulfuration and hydrogen sulfide production

ABSTRACTCysteine desulfuration is one of the main ways for hydrogen sulfide (H2S) generation in cells and is usually conducted by cystathionine γ-lyase. Herein, we describe a newly discovered deep-sea bacterial threonine dehydratase (psTD), which is surprisingly discovered to drive L-cysteine desulfuration. The mechanisms of psTD catalyzing cysteine desulfuration towards H2S production are first clarified in vitro and in vivo through a combination of genetic and biochemical methods. Furthermore, based on the solved structures of psTD and its various mutants, two or three pockets are found in the active site of psTD, and switch states between inward and outward orientation of a key amino acid R77 determine the open or close status of Pocket III for small molecule exchanges, which further facilitates cysteine desulfuration. Our results reveal the functional diversity and structural specificity of psTD towards L-cysteine desulfuration and H2S formation. Given the broad distribution of psTD homologs in different bacteria, we speculate that some threonine dehydratases have evolved a novel function towards cysteine desulfuration, which benefits the producer to utilize cysteine as a sulfur source for better adapting external environments.

Identification of an l-serine/l-threonine dehydratase with glutamate racemase activity in mammals

Recent investigations have shown that multiple d-amino acids are present in mammals and these compounds have distinctive physiological functions. Free d-glutamate is present in various mammalian tissues and cells and in particular, it is presumably correlated with cardiac function, and much interest is growing in its unique metabolic pathways. Recently, we first identified d-glutamate cyclase as its degradative enzyme in mammals, whereas its biosynthetic pathway in mammals is unclear. Glutamate racemase is a most probable candidate, which catalyzes interconversion between d-glutamate and l-glutamate. Here, we identified the cDNA encoding l-serine dehydratase-like (SDHL) as the first mammalian clone with glutamate racemase activity. This rat SDHL had been deposited in mammalian databases as a protein of unknown function and its amino acid sequence shares ∼60% identity with that of l-serine dehydratase. Rat SDHL was expressed in Escherichia coli, and the enzymatic properties of the recombinant were characterized. The results indicated that rat SDHL is a multifunctional enzyme with glutamate racemase activity in addition to l-serine/l-threonine dehydratase activity. This clone is hence abbreviated as STDHgr. Further experiments using cultured mammalian cells confirmed that d-glutamate was synthesized and l-serine and l-threonine were decomposed. It was also found that SDHL (STDHgr) contributes to the homeostasis of several other amino acids.

Threonine in mammals: indispensability and transamination

As is known, amino acid threonine is not synthesized in the vertebrates when it does not come with food and the decomposition of threonine under the action of threonine dehydratase is irreversible process. Some facts point to the presence of insignificant threonine synthesis in animals. The question arises about the possibility of biosynthesis of threonine in animals in the absence of it in food, that is, its interchangeability. Research on this issue is important for compiling the diet of animals. The article shows that the threonine cannot be synthesized by reversibility of the reaction of its decomposition as well why threonine dehydrogenase in the tissues of mammals cannot be used in threonine biosynthesis. It is concluded that some quantity of threonine is involved in transamination.

Co-Expression of Threonine Dehydratase and Acetolactate Synthase in Escherichia Coli for L-Isoleucine Production

Metabolic engineering ofCorynebacterium glutamicumhas sought to divert carbon into L-isoleucine. However, the fermentation period of this strain is long. TheC.glutamicumYILW strain (LeuL, AHVr, SGr, Leu-MEr) was previously derived by repeated compound mutagenesis which could accumulate 20.2 g/L L-isoleucine in a 5-L jar fermentor. Overexpression of the threonine dehydratase gene (ilvA) fromCorynebacterium glutamicumYILW and coexpression of threonine dehydratase and acetolactate synthase (ilvBN) from it were employed to divert carbon flux toward L-isoleucine. The strainE. coliTRFC with the expression ofilvA could accumulate L-isoleucine of 6.8 g/L without accumulation of any L-threonine by fed-batch fermentation in a 5-L jar fermentor. However, the production of L-isoleucine by the strainE.coliTRFC with the co-expression ofilvA andilvBN was decreased by 19.1%, and the production of L-valine was increased by 40% compared with that ofE. coliTRFC with the expression ofilvA.