Your browser doesn't support javascript.
loading
Direct evidence that an extended hydrogen-bonding network influences activation of pyridoxal 5'-phosphate in aspartate aminotransferase.
Dajnowicz, Steven; Parks, Jerry M; Hu, Xiche; Gesler, Korie; Kovalevsky, Andrey Y; Mueser, Timothy C.
Afiliación
  • Dajnowicz S; From the Department of Chemistry and Biochemistry, University of Toledo, Toledo, Ohio 43606.
  • Parks JM; the Biology and Soft Matter Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee 37831, and.
  • Hu X; the University of Tennessee/Oak Ridge National Laboratory Center for Molecular Biophysics, Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee 37831.
  • Gesler K; From the Department of Chemistry and Biochemistry, University of Toledo, Toledo, Ohio 43606.
  • Kovalevsky AY; From the Department of Chemistry and Biochemistry, University of Toledo, Toledo, Ohio 43606.
  • Mueser TC; the Biology and Soft Matter Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee 37831, and.
J Biol Chem ; 292(14): 5970-5980, 2017 04 07.
Article en En | MEDLINE | ID: mdl-28232482
Pyridoxal 5'-phosphate (PLP) is a fundamental, multifunctional enzyme cofactor used to catalyze a wide variety of chemical reactions involved in amino acid metabolism. PLP-dependent enzymes optimize specific chemical reactions by modulating the electronic states of PLP through distinct active site environments. In aspartate aminotransferase (AAT), an extended hydrogen bond network is coupled to the pyridinyl nitrogen of the PLP, influencing the electrophilicity of the cofactor. This network, which involves residues Asp-222, His-143, Thr-139, His-189, and structural waters, is located at the edge of PLP opposite the reactive Schiff base. We demonstrate that this hydrogen bond network directly influences the protonation state of the pyridine nitrogen of PLP, which affects the rates of catalysis. We analyzed perturbations caused by single- and double-mutant variants using steady-state kinetics, high resolution X-ray crystallography, and quantum chemical calculations. Protonation of the pyridinyl nitrogen to form a pyridinium cation induces electronic delocalization in the PLP, which correlates with the enhancement in catalytic rate in AAT. Thus, PLP activation is controlled by the proximity of the pyridinyl nitrogen to the hydrogen bond microenvironment. Quantum chemical calculations indicate that Asp-222, which is directly coupled to the pyridinyl nitrogen, increases the pKa of the pyridine nitrogen and stabilizes the pyridinium cation. His-143 and His-189 also increase the pKa of the pyridine nitrogen but, more significantly, influence the position of the proton that resides between Asp-222 and the pyridinyl nitrogen. These findings indicate that the second shell residues directly enhance the rate of catalysis in AAT.
Asunto(s)
Palabras clave
Texto completo
Añadir a Mi BVS
Imprimir
XML
PubMed Links
Buscar en Google

Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Fosfato de Piridoxal / Aspartato Aminotransferasas / Modelos Moleculares Tipo de estudio: Prognostic_studies Límite: Animals Idioma: En Revista: J Biol Chem Año: 2017 Tipo del documento: Article Pais de publicación: Estados Unidos
Texto completo
Añadir a Mi BVS
Imprimir
XML
PubMed Links
Buscar en Google

Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Fosfato de Piridoxal / Aspartato Aminotransferasas / Modelos Moleculares Tipo de estudio: Prognostic_studies Límite: Animals Idioma: En Revista: J Biol Chem Año: 2017 Tipo del documento: Article Pais de publicación: Estados Unidos