Heterogeneity of active sites in recombinant betaine aldehyde dehydrogenase is modulated by potassium. 2020

César Muñoz-Bacasehua, and Jesús A Rosas-Rodríguez, and Aldo A Arvizu-Flores, and Aurora Stephens-Camacho, and José G Soñanez-Organis, and Ciria G Figueroa-Soto, and Elisa M Valenzuela-Soto
Ciencia de los Alimentos, Centro de Investigación en Alimentación y Desarrollo A.C., Hermosillo, México.

Betaine aldehyde dehydrogenase (BADH EC 1.2.1.8) catalyzes the irreversible oxidation of betaine aldehyde to glycine betaine using NAD+ as a coenzyme. Porcine kidney BADH (pkBADH) follows a bi-bi ordered mechanism in which NAD+ binds to the enzyme before the aldehyde. Previous studies showed that NAD+ induces complex and unusual conformational changes on pkBADH and that potassium is required to maintain its quaternary structure. The aim of this work was to analyze the structural changes in pkBADH caused by NAD+ binding and the role played by potassium in those changes. The pkBADH cDNA was cloned and overexpressed in Escherichia coli, and the protein was purified by affinity chromatography using a chitin matrix. The pkBADH/NAD+ interaction was analyzed by circular dichroism (CD) and by isothermal titration calorimetry (ITC) by titrating the enzyme with NAD+ . The cDNA has an open reading frame of 1485 bp and encodes a protein of 494 amino acids, with a predicted molecular mass of 53.9 kDa. CD data showed that the binding of NAD+ to the enzyme caused changes in its secondary structure, whereas the presence of K+ helps maintain its α-helix content. K+ increased the thermal stability of the pkBADH-NAD+ complex by 5.3°C. ITC data showed that NAD+ binding occurs with different association constants for each active site between 37.5 and 8.6 μM. All the results support previous data in which the enzyme incubation with NAD+ provoked changes in reactivity, which is an indication of slow conformational rearrangements of the active site.

UI MeSH Term Description Entries
D007668 Kidney Body organ that filters blood for the secretion of URINE and that regulates ion concentrations. Kidneys
D011188 Potassium An element in the alkali group of metals with an atomic symbol K, atomic number 19, and atomic weight 39.10. It is the chief cation in the intracellular fluid of muscle and other cells. Potassium ion is a strong electrolyte that plays a significant role in the regulation of fluid volume and maintenance of the WATER-ELECTROLYTE BALANCE.
D011487 Protein Conformation The characteristic 3-dimensional shape of a protein, including the secondary, supersecondary (motifs), tertiary (domains) and quaternary structure of the peptide chain. PROTEIN STRUCTURE, QUATERNARY describes the conformation assumed by multimeric proteins (aggregates of more than one polypeptide chain). Conformation, Protein,Conformations, Protein,Protein Conformations
D006863 Hydrogen-Ion Concentration The normality of a solution with respect to HYDROGEN ions; H+. It is related to acidity measurements in most cases by pH pH,Concentration, Hydrogen-Ion,Concentrations, Hydrogen-Ion,Hydrogen Ion Concentration,Hydrogen-Ion Concentrations
D000595 Amino Acid Sequence The order of amino acids as they occur in a polypeptide chain. This is referred to as the primary structure of proteins. It is of fundamental importance in determining PROTEIN CONFORMATION. Protein Structure, Primary,Amino Acid Sequences,Sequence, Amino Acid,Sequences, Amino Acid,Primary Protein Structure,Primary Protein Structures,Protein Structures, Primary,Structure, Primary Protein,Structures, Primary Protein
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D013696 Temperature The property of objects that determines the direction of heat flow when they are placed in direct thermal contact. The temperature is the energy of microscopic motions (vibrational and translational) of the particles of atoms. Temperatures
D016415 Sequence Alignment The arrangement of two or more amino acid or base sequences from an organism or organisms in such a way as to align areas of the sequences sharing common properties. The degree of relatedness or homology between the sequences is predicted computationally or statistically based on weights assigned to the elements aligned between the sequences. This in turn can serve as a potential indicator of the genetic relatedness between the organisms. Sequence Homology Determination,Determination, Sequence Homology,Alignment, Sequence,Alignments, Sequence,Determinations, Sequence Homology,Sequence Alignments,Sequence Homology Determinations
D050728 Betaine-Aldehyde Dehydrogenase An NAD+ dependent enzyme that catalyzes the oxidation of betain aldehyde to BETAINE. Betaine Aldehyde Dehydrogenase,Betaine Aldehyde Oxidase,Glycine Betaine Aldehyde Dehydrogenase,Aldehyde Dehydrogenase, Betaine,Aldehyde Oxidase, Betaine,Dehydrogenase, Betaine Aldehyde,Dehydrogenase, Betaine-Aldehyde,Oxidase, Betaine Aldehyde
D020134 Catalytic Domain The region of an enzyme that interacts with its substrate to cause the enzymatic reaction. Active Site,Catalytic Core,Catalytic Region,Catalytic Site,Catalytic Subunit,Reactive Site,Active Sites,Catalytic Cores,Catalytic Domains,Catalytic Regions,Catalytic Sites,Catalytic Subunits,Core, Catalytic,Cores, Catalytic,Domain, Catalytic,Domains, Catalytic,Reactive Sites,Region, Catalytic,Regions, Catalytic,Site, Active,Site, Catalytic,Site, Reactive,Sites, Active,Sites, Catalytic,Sites, Reactive,Subunit, Catalytic,Subunits, Catalytic

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