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Structural Basis for Recognition of L-lysine, L-ornithine, and L-2,4-diamino Butyric Acid by Lysine Cyclodeaminase.

ABSTRACT: L-pipecolic acid is a non-protein amino acid commonly found in plants, animals, and microorganisms. It is a well-known precursor to numerous microbial secondary metabolites and pharmaceuticals, including anticancer agents, immunosuppressants, and several antibiotics. Lysine cyclodeaminase (LCD) catalyzes ?-deamination of L-lysine into L-pipecolic acid using ?-nicotinamide adenine dinucleotide as a cofactor. Expression of a human homolog of LCD, ?-crystallin, is elevated in prostate cancer patients. To understand the structural features and catalytic mechanisms of LCD, we determined the crystal structures of Streptomyces pristinaespiralis LCD (SpLCD) in (i) a binary complex with NAD+, (ii) a ternary complex with NAD+ and L-pipecolic acid, (iii) a ternary complex with NAD+ and L-proline, and (iv) a ternary complex with NAD+ and L-2,4-diamino butyric acid. The overall structure of SpLCD was similar to that of ornithine cyclodeaminase from Pseudomonas putida. In addition, SpLCD recognized L-lysine, L-ornithine, and L-2,4-diamino butyric acid despite differences in the active site, including differences in hydrogen bonding by Asp236, which corresponds with Asp228 from Pseudomonas putida ornithine cyclodeaminase. The substrate binding pocket of SpLCD allowed substrates smaller than lysine to bind, thus enabling binding to ornithine and L-2,4-diamino butyric acid. Our structural and biochemical data facilitate a detailed understanding of substrate and product recognition, thus providing evidence for a reaction mechanism for SpLCD. The proposed mechanism is unusual in that NAD+ is initially converted into NADH and then reverted back into NAD+ at a late stage of the reaction.

SUBMITTER: Min K

PROVIDER: S-EPMC5935100 | biostudies-literature | 2018 Apr

REPOSITORIES: biostudies-literature

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Structural Basis for Recognition of L-lysine, L-ornithine, and L-2,4-diamino Butyric Acid by Lysine Cyclodeaminase.

Min Kyungjin K Yoon Hye-Jin HJ Matsuura Atsushi A Kim Yong Hwan YH Lee Hyung Ho HH

Molecules and cells 20180405 4

L-pipecolic acid is a non-protein amino acid commonly found in plants, animals, and microorganisms. It is a well-known precursor to numerous microbial secondary metabolites and pharmaceuticals, including anticancer agents, immunosuppressants, and several antibiotics. Lysine cyclodeaminase (LCD) catalyzes β-deamination of L-lysine into L-pipecolic acid using β-nicotinamide adenine dinucleotide as a cofactor. Expression of a human homolog of LCD, μ-crystallin, is elevated in prostate cancer patien ...[more]

PMID: 29629557

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Project description:BackgroundIn plants, proline synthesis occurs by two enzymatic steps starting from glutamate as a precursor. Some bacteria, including bacteria such as Agrobacterium rhizogenes have an Ornithine Cyclodeaminase (OCD) which can synthesize proline in a single step by deamination of ornithine. In A. rhizogenes, OCD is one of the genes transferred to the plant genome during the transformation process and plants expressing A. rhizogenes OCD have developmental phenotypes. One nuclear encoded gene of Arabidopsis thaliana has recently been annotated as an OCD (OCD-like; referred to here as AtOCD) but nothing is known of its function. As proline metabolism contributes to tolerance of low water potential during drought, it is of interest to determine if AtOCD affects proline accumulation or low water potential tolerance.ResultsExpression of AtOCD was induced by low water potential stress and by exogenous proline, but not by the putative substrate ornithine. The AtOCD protein was plastid localized. T-DNA mutants of atocd and AtOCD RNAi plants had approximately 15% higher proline accumulation at low water potential while p5cs1-4/atocd double mutants had 40% higher proline than p5cs1 at low water potential but no change in proline metabolism gene expression which could directly explain the higher proline level. AtOCD overexpression did not affect proline accumulation. Enzymatic assays with bacterially expressed AtOCD or AtOCD purified from AtOCD:Flag transgenic plants did not detect any activity using ornithine, proline or several other amino acids as substrates. Moreover, AtOCD mutant or over-expression lines had normal morphology and no difference in root elongation or flowering time, in contrast to previous report of transgenic plants expressing A. rhizogenes OCD. Metabolite analysis found few differences between AtOCD mutants and overexpression lines.ConclusionsThe Arabidopsis OCD-like protein (AtOCD) may not catalyze ornithine to proline conversion and this is consistent with observation that three residues critical for activity of bacterial OCDs are not conserved in AtOCD. AtOCD was, however, stress and proline induced and lack of AtOCD expression increased proline accumulation by an unknown mechanism which did not require expression of P5CS1, the main enzyme responsible for stress-induced proline synthesis from glutamate. The results suggest that AtOCD may have function distinct from bacterial OCDs.

Dataset Information

Structural Basis for Recognition of L-lysine, L-ornithine, and L-2,4-diamino Butyric Acid by Lysine Cyclodeaminase.

Publications

Structural Basis for Recognition of L-lysine, L-ornithine, and L-2,4-diamino Butyric Acid by Lysine Cyclodeaminase.

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