5B55

Crystal structure of hydrogen sulfide-producing enzyme (Fn1055) D232N mutant in complexed with alpha-aminoacrylate intermediate: lysine-dimethylated form

Summary for 5B55

• Entry DOI: 10.2210/pdb5b55/pdb
• Related: 5B53 5B54
• Descriptor: Cysteine synthase, PYRIDOXAL-5'-PHOSPHATE, 2-{[(E)-{3-hydroxy-2-methyl-5-[(phosphonooxy)methyl]pyridin-4-yl}methylidene]amino}prop-2-enoic acid, ... (5 entities in total)
• Functional Keywords: pyridoxal 5'-phosphate dependent enzyme, alpha-aminoacrylate, transferase
• Biological source: Fusobacterium nucleatum subsp. nucleatum strain ATCC 25586
• Total number of polymer chains: 2
• Total formula weight: 78467.16

Authors

Kezuka, Y.,Yoshida, Y.,Nonaka, T. (deposition date: 2016-04-22, release date: 2017-04-26, Last modification date: 2023-11-08)

Primary citation

Kezuka, Y.,Ishida, T.,Yoshida, Y.,Nonaka, T.
Structural insights into the catalytic mechanism of cysteine (hydroxyl) lyase from the hydrogen sulfide-producing oral pathogen,Fusobacterium nucleatum.
Biochem. J., 475:733-748, 2018

PubMed Abstract: Hydrogen sulfide (HS) plays important roles in the pathogenesis of periodontitis. Oral pathogens typically produce HS from l-cysteine in addition to pyruvate and [Formula: see text] However, from subsp. ATCC 25586 encodes a pyridoxal 5'-phosphate (PLP)-dependent enzyme that catalyzes the production of HS and l-serine from l-cysteine and HO, an unusual cysteine (hydroxyl) lyase reaction (β-replacement reaction). To reveal the reaction mechanism, the crystal structure of substrate-free Fn1055 was determined. Based on this structure, a model of the l-cysteine-PLP Schiff base suggested that the thiol group forms hydrogen bonds with Asp and Ser, and the substrate α-carboxylate interacts with Thr and Gln Asp is a unique residue to Fn1055 and its substitution to asparagine (D232N) resulted in almost complete loss of β-replacement activity. The D232N structure obtained in the presence of l-cysteine contained the α-aminoacrylate-PLP Schiff base in the active site, indicating that Asp is essential for the addition of water to the α-aminoacrylate to produce the l-serine-PLP Schiff base. Rapid-scan stopped-flow kinetic analyses showed an accumulation of the α-aminoacrylate intermediate during the reaction cycle, suggesting that water addition mediated by Asp is the rate-limiting step. In contrast, mutants containing substitutions of other active-site residues (Ser, Thr, and Gln) exhibited reduced β-replacement activity by more than 100-fold. Finally, based on the structural and biochemical analyses, we propose a mechanism of the cysteine (hydroxyl) lyase reaction by Fn1055. The present study leads to elucidation of the HS-producing mechanism in .

PubMed: 29343611
DOI: 10.1042/BCJ20170838

Experimental method

X-RAY DIFFRACTION (2.14 Å)