6ojw | pdb_00006ojw
Crystal structure of Sphingomonas paucimobilis TMY1009 holo-LsdA
Structural highlights
Function[LSDX1_SPHPI] Catalyzes the cleavage of the interphenyl double bond (C alpha-C beta) of lignin-derived polyphenolic diaryl-propane type compounds (Stilbene).[1] [2] [3] [4] Publication Abstract from PubMedLignostilbene-alpha,beta-dioxygenase A (LsdA) from the bacterium Sphingomonas paucimobilis TMY1009 is a non-heme iron oxygenase that catalyzes the cleavage of lignostilbene, a compound arising in lignin transformation, to two vanillin molecules. To examine LsdA's substrate specificity, we heterologously produced the dimeric enzyme with the help of chaperones. When tested on several substituted stilbenes, LsdA exhibited greatest specificity for lignostilbene (k cat (app)/K M (app)= 1.00 +/- 0.04 x 10(6) M(-1)s(-1)). These experiments further indicated that the substrate's 4-hydroxy moiety is required for catalysis, and that this moiety cannot be replaced with a methoxy group. Phenylazophenol inhibited the LsdA-catalyzed cleavage of lignostilbene in a reversible, mixed fashion (K ic = 6 +/- 1 microM, Kiu = 24 +/- 4 microM). An X-ray crystal structure of LsdA at 2.3 A resolution revealed a seven-bladed beta-propeller fold with an iron cofactor coordinated by four histidines, in agreement with previous observations on related carotenoid cleavage oxygenases. We noted that residues at the dimer interface are also present in LsdB, another lignostilbene dioxygenase in S. paucimobilis TMY1009, rationalizing LsdA and LsdB homo- and heterodimerization in vivo A structure of a LsdA.phenylazophenol complex identified Phe-59, Tyr-101, and Lys-134 as contacting the 4-hydroxyphenyl moiety of the inhibitor. Phe-59 and Tyr-101 substitutions with His and Phe, respectively, reduced LsdA activity (k cat (app)) ~15- and 10-fold. The K134M variant did not detectably cleave lignostilbene, indicating that Lys-134 plays a key catalytic role. This study expands our mechanistic understanding of LsdA and related stilbene-cleaving dioxygenases. Identification of functionally important residues and structural features in a bacterial lignostilbene dioxygenase.,Kuatsjah E, Verstraete MM, Kobylarz MJ, Liu AKN, Murphy MEP, Eltis LD J Biol Chem. 2019 Jul 10. pii: RA119.009428. doi: 10.1074/jbc.RA119.009428. PMID:31292192[5] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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