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Comparative structural and enzymatic studies on Salmonella typhimurium diaminopropionate ammonia lyase reveal its unique features

Deka, G and Bisht, S and Savithri, HS and Murthy, MRN (2018) Comparative structural and enzymatic studies on Salmonella typhimurium diaminopropionate ammonia lyase reveal its unique features. In: Journal of Structural Biology, 202 (2). pp. 118-128.

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Official URL: https://doi.org/10.1016/j.jsb.2017.12.012


Cellular metabolism of amino acids is controlled by a large number of pyridoxal 5′-phosphate (PLP) dependent enzymes. Diaminopropionate ammonia lyase (DAPAL), a fold type II PLP-dependent enzyme, degrades both the D and L forms of diaminopropionic acid (DAP) to pyruvate and ammonia. Earlier studies on the Escherichia coli DAPAL (EcDAPAL) had suggested that a disulfide bond located close to the active site may be crucial for maintaining the geometry of the substrate entry channel and the active site. In order to obtain further insights into the catalytic properties of DAPAL, structural and functional studies on Salmonella typhimurium DAPAL (StDAPAL) were initiated. The three-dimensional X-ray crystal structure of StDAPAL was determined at 2.5 Å resolution. As expected, the polypeptide fold and dimeric organization of StDAPAL is similar to those of EcDAPAL. A phosphate group was located in the active site of StDAPAL and expulsion of this phosphate is probably essential to bring Asp125 to a conformation suitable for proton abstraction from the substrate (D-DAP). The unique disulfide bond of EcDAPAL was absent in StDAPAL, although the enzyme displayed comparable catalytic activity. Site directed mutagenesis of the cysteine residues involved in disulfide bond formation in EcDAPAL followed by functional and biophysical studies further confirmed that the disulfide bond is not necessary either for substrate binding or for catalysis. The activity of StDAPAL but not EcDAPAL was enhanced by monovalent cations suggesting subtle differences in the active site geometries of these two closely related enzymes.

Item Type: Journal Article
Publication: Journal of Structural Biology
Publisher: Academic Press Inc.
Additional Information: The copyright for this article belongs to the Academic Press Inc.
Keywords: aspartic acid; cysteine; diaminopropionate ammonia lyase; lyase; phosphate; unclassified drug; ammonia lyase; diaminopropionate ammonia-lyase, Article; biophysics; catalysis; controlled study; crystal structure; disulfide bond; enzyme active site; enzyme activity; enzyme analysis; enzyme conformation; enzyme structure; enzyme substrate complex; nonhuman; priority journal; Salmonella enterica serovar Typhimurium; site directed mutagenesis; X ray diffraction; chemistry; enzyme specificity; enzymology; Escherichia coli; genetics; kinetics; protein folding; Salmonella enterica serovar Typhimurium; structure activity relation; X ray crystallography, Ammonia-Lyases; Catalysis; Catalytic Domain; Crystallography, X-Ray; Escherichia coli; Kinetics; Mutagenesis, Site-Directed; Protein Folding; Salmonella typhimurium; Structure-Activity Relationship; Substrate Specificity
Department/Centre: Division of Biological Sciences > Biochemistry
Division of Biological Sciences > Molecular Biophysics Unit
Date Deposited: 08 Aug 2022 06:26
Last Modified: 08 Aug 2022 06:26
URI: https://eprints.iisc.ac.in/id/eprint/75568

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