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Analysis of the Active Site Cysteine Residue of the Sacrificial Sulfur Insertase LarE from Lactobacillus plantarum.
Fellner, Matthias; Rankin, Joel A; Desguin, Benoît; Hu, Jian; Hausinger, Robert P.
Afiliación
  • Fellner M; Department of Biochemistry and Molecular Biology , Michigan State University , East Lansing , Michigan 48824 , United States.
  • Rankin JA; Department of Biochemistry and Molecular Biology , Michigan State University , East Lansing , Michigan 48824 , United States.
  • Desguin B; Institute of Life Sciences , Université catholique de Louvain , B-1348 Louvain-La-Neuve , Belgium.
  • Hu J; Department of Biochemistry and Molecular Biology , Michigan State University , East Lansing , Michigan 48824 , United States.
  • Hausinger RP; Department of Chemistry , Michigan State University , East Lansing , Michigan 48824 , United States.
Biochemistry ; 57(38): 5513-5523, 2018 09 25.
Article en En | MEDLINE | ID: mdl-30157639
LarE from Lactobacillus plantarum is an ATP-dependent sulfur transferase that sacrifices its Cys176 sulfur atom to form a dehydroalanine (Dha) side chain during biosynthesis of the covalently linked nickel-pincer nucleotide (NPN) cofactor (pyridinium 3-thioamide-5-thiocarboxylic acid mononucleotide) of lactate racemase. Coenzyme A (CoA) stabilizes LarE and forms a CoA-Cys176 mixed disulfide with the protein. This study presents the crystal structure of the LarE/CoA complex, revealing protein interactions with CoA that mimic those for binding ATP. CoA weakly inhibits LarE activity, and the persulfide of CoA is capable of partially regenerating functional LarE from the Dha176 form of the protein. The physiological relevance of this cycling reaction is unclear. A new form of LarE was discovered, an NPN-LarE covalent adduct, explaining prior results in which activation of the lactate racemase apoprotein required only the isolated LarE. The crystal structure of the inactive C176A variant revealed a fold essentially identical to that of wild-type LarE. Additional active site variants of LarE were created and their activities characterized, with all LarE variants analyzed in terms of the structure. Finally, the L. plantarum LarE structure was compared to a homology model of Thermoanaerobacterium thermosaccharolyticum LarE, predicted to contain three cysteine residues at the active site, and to other proteins with a similar fold and multiple active site cysteine residues. These findings suggest that some LarE orthologs may not be sacrificial but instead may catalyze sulfur transfer by using a persulfide mechanism or from a labile site on a [4Fe-4S] cluster at this position.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Azufre / Proteínas Bacterianas / Racemasas y Epimerasas / Cisteína / Lactobacillus plantarum Tipo de estudio: Prognostic_studies Idioma: En Revista: Biochemistry Año: 2018 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos
Texto completo
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XML
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Azufre / Proteínas Bacterianas / Racemasas y Epimerasas / Cisteína / Lactobacillus plantarum Tipo de estudio: Prognostic_studies Idioma: En Revista: Biochemistry Año: 2018 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos