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Structural basis for catalytic racemization and substrate specificity of an N-acylamino acid racemase homologue from Deinococcus radiodurans.

Wang WC, Chiu WC, Hsu SK, Wu CL, Chen CY, Liu JS, Hsu WH
J Mol Biol 342 155-69 (2004)
Related entries: 1r0m, 1xs2

Cited: 10 times
EuropePMC logo PMID: 15313614

Abstract

N-acylamino acid racemase (NAAAR) catalyzes the racemization of N-acylamino acids and can be used in concert with an aminoacylase to produce enantiopure alpha-amino acids, a process that has potential industrial applications. Here we have cloned and characterized an NAAAR homologue from a radiation-resistant ancient bacterium, Deinococcus radiodurans. The expressed NAAAR racemized various substrates at an optimal temperature of 60 degrees C and had Km values of 24.8 mM and 12.3 mM for N-acetyl-D-methionine and N-acetyl-L-methionine, respectively. The crystal structure of NAAAR was solved to 1.3 A resolution using multiwavelength anomalous dispersion (MAD) methods. The structure consists of a homooctamer in which each subunit has an architecture characteristic of enolases with a capping domain and a (beta/alpha)7 beta barrel domain. The NAAAR.Mg2+ and NAAAR.N-acetyl-L-glutamine.Mg2+ structures were also determined, allowing us to define the Lys170-Asp195-Glu220-Asp245-Lys269 framework for catalyzing 1,1-proton exchange of N-acylamino acids. Four subsites enclosing the substrate are identified: catalytic site, metal-binding site, side-chain-binding region, and a flexible lid region. The high conservation of catalytic and metal-binding sites in different enolases reflects the essentiality of a common catalytic platform, allowing these enzymes to robustly abstract alpha-protons of various carboxylate substrates efficiently. The other subsites involved in substrate recognition are less conserved, suggesting that divergent evolution has led to functionally distinct enzymes.

Articles - 1xpy mentioned but not cited (1)

  1. Comparison of Alicyclobacillus acidocaldarius o-Succinylbenzoate Synthase to Its Promiscuous N-Succinylamino Acid Racemase/ o-Succinylbenzoate Synthase Relatives. Odokonyero D, McMillan AW, Ramagopal UA, Toro R, Truong DP, Zhu M, Lopez MS, Somiari B, Herman M, Aziz A, Bonanno JB, Hull KG, Burley SK, Romo D, Almo SC, Glasner ME. Biochemistry 57 3676-3689 (2018)


Articles citing this publication (9)

  1. Unusual arginine formations in protein function and assembly: rings, strings, and stacks. Neves MA, Yeager M, Abagyan R. J Phys Chem B 116 7006-7013 (2012)
  2. Structure-stability-activity relationship in covalently cross-linked N-carbamoyl D-amino acid amidohydrolase and N-acylamino acid racemase. Chiu WC, You JY, Liu JS, Hsu SK, Hsu WH, Shih CH, Hwang JK, Wang WC. J Mol Biol 359 741-753 (2006)
  3. Isolation and characterization of a novel lysine racemase from a soil metagenomic library. Chen IC, Lin WD, Hsu SK, Thiruvengadam V, Hsu WH. Appl Environ Microbiol 75 5161-5166 (2009)
  4. Loss of quaternary structure is associated with rapid sequence divergence in the OSBS family. Odokonyero D, Sakai A, Patskovsky Y, Malashkevich VN, Fedorov AA, Bonanno JB, Fedorov EV, Toro R, Agarwal R, Wang C, Ozerova ND, Yew WS, Sauder JM, Swaminathan S, Burley SK, Almo SC, Glasner ME. Proc Natl Acad Sci U S A 111 8535-8540 (2014)
  5. Structure and conformational stability of a tetrameric thermostable N-succinylamino acid racemase. Pozo-Dengra J, Martínez-Rodríguez S, Contreras LM, Prieto J, Andújar-Sánchez M, Clemente-Jiménez JM, Las Heras-Vázquez FJ, Rodríguez-Vico F, Neira JL. Biopolymers 91 757-772 (2009)
  6. Enantioselective synthesis of L-homophenylalanine by whole cells of recombinant Escherichia coli expressing L-aminoacylase and N-acylamino acid racemase genes from Deinococcus radiodurans BCRC12827. Hsu SK, Lo HH, Kao CH, Lee DS, Hsu WH. Biotechnol Prog 22 1578-1584 (2006)
  7. Promiscuity of Exiguobacterium sp. AT1b o-succinylbenzoate synthase illustrates evolutionary transitions in the OSBS family. Brizendine AM, Odokonyero D, McMillan AW, Zhu M, Hull K, Romo D, Glasner ME. Biochem Biophys Res Commun 450 679-684 (2014)
  8. Crystal structure of N-acylamino acid racemase from Thermus thermophilus HB8. Hayashida M, Kim SH, Takeda K, Hisano T, Miki K. Proteins 71 519-523 (2008)
  9. Biochemical and Mutational Characterization of N-Succinyl-Amino Acid Racemase from Geobacillus stearothermophilus CECT49. Soriano-Maldonado P, Andújar-Sánchez M, Clemente-Jiménez JM, Rodríguez-Vico F, Las Heras-Vázquez FJ, Martínez-Rodríguez S. Mol Biotechnol 57 454-465 (2015)


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