Structural highlights
Function[A0A0U3H6N8_9CREN] Involved in the biosynthesis of branched-chain amino acids (BCAA). Catalyzes an alkyl-migration followed by a ketol-acid reduction of (S)-2-acetolactate (S2AL) to yield (R)-2,3-dihydroxy-isovalerate. In the isomerase reaction, S2AL is rearranged via a Mg-dependent methyl migration to produce 3-hydroxy-3-methyl-2-ketobutyrate (HMKB). In the reductase reaction, this 2-ketoacid undergoes a metal-dependent reduction by NADPH to yield (R)-2,3-dihydroxy-isovalerate.[HAMAP-Rule:MF_00435] Publication Abstract from PubMedKetol-acid reductoisomerase (KARI) is a bifunctional enzyme in the second step of branched-chain amino acids biosynthetic pathway. Most KARIs prefer NADPH as a cofactor. However, KARI with a preference for NADH is desirable in industrial applications including anaerobic fermentation for the production of branched-chain amino acids or biofuels. Here, we characterize a thermoacidophilic archaeal Sac-KARI from Sulfolobus acidocaldarius and present its crystal structure at a 1.75-A resolution. By comparison with other holo-KARI structures, one sulphate ion is observed in each binding site for the 2'-phosphate of NADPH, implicating its NADPH preference. Sac-KARI has very high affinity for NADPH and NADH, with K M values of 0.4 muM for NADPH and 6.0 muM for NADH, suggesting that both are good cofactors at low concentrations although NADPH is favoured over NADH. Furthermore, Sac-KARI can catalyze 2(S)-acetolactate (2S-AL) with either cofactor from 25 to 60 degrees C, but the enzyme has higher activity by using NADPH. In addition, the catalytic activity of Sac-KARI increases significantly with elevated temperatures and reaches an optimum at 60 degrees C. Bi-cofactor utilization and the thermoactivity of Sac-KARI make it a potential candidate for use in metabolic engineering or industrial applications under anaerobic or harsh conditions. NADH/NADPH bi-cofactor-utilizing and thermoactive ketol-acid reductoisomerase from Sulfolobus acidocaldarius.,Chen CY, Ko TP, Lin KF, Lin BL, Huang CH, Chiang CH, Horng JC Sci Rep. 2018 May 8;8(1):7176. doi: 10.1038/s41598-018-25361-4. PMID:29739976[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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