PL EN


Preferences help
enabled [disable] Abstract
Number of results
2015 | 62 | 4 | 903-912
Article title

Characterization of two aminotransferases from Candida albicans

Content
Title variants
Languages of publication
EN
Abstracts
EN
Aminoadipate aminotransferase (AmAA) is an enzyme of α-aminoadipate pathway (AAP) for l-lysine biosynthesis. AmAA may also participated in biosynthesis or degradation of aromatic amino acids and in d-tryptophan based pigment production. The AAP is unique for fungal microorganisms. Enzymes involved in this pathway have specific structures and properties. These features can be used as potential molecular markers. Enzymes catalyzing reactions of l-lysine biosynthesis in Candida albicans may also become new targets for antifungal chemotherapy. Search of the NCBI database resulted in identification of two putative aminoadipate aminotransferase genes from Candida albicans: ARO8 (ORFs 19.2098 and 19.9645) and YER152C (ORFs 19.1180 and 19.8771). ARO8 from C. albicans exhibits 53% identity to ARO8 from S. cerevisiae, while YER152C exhibits 30% identity to ARO8 and 45% to YER152C from S. cerevisiae. We amplified two genes from the C. albicans genome: ARO8 and YER152C. Both were cloned and expressed as His-tagged fusion proteins in E. coli. The purified Aro8CHp gene product revealed aromatic and α-aminoadipate aminotransferase activity. Basic molecular properties of the purified protein were determined. We obtained catalytic parameters of Aro8CHp with aromatic amino acids and aminoadipate (AA) (Km(L-Phe) 0.05±0.003 mM, Km(L-Tyr) 0.1±0.008 mM, Km(L-AA) 0.02±0.006 mM) and confirmed the enzyme broad substrate spectrum. The assays also demonstrated that this enzyme may use 2-oxoadipate and 2-oxoglutarate (2-OG) as amino acceptors. Aro8-CHp exhibited pH optima range of 8, which is similar to AmAA from S. cerevisiae. Our results also indicate that CaYer152Cp has a possible role only in aromatic amino acids degradation, in contrast to CaAro8CHp.
Publisher

Year
Volume
62
Issue
4
Pages
903-912
Physical description
Dates
published
2015
received
2015-07-31
revised
2015-10-09
accepted
2015-10-29
(unknown)
2015-11-30
Contributors
author
  • Department of Pharmaceutical Technology and Biochemistry, Gdansk University of Technology, Gdańsk, Poland
author
  • Department of Pharmaceutical Technology and Biochemistry, Gdansk University of Technology, Gdańsk, Poland
References
  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248-254.
  • Brunke S, Seider K, Almeida RS, Heyken A, Fleck CB, Brock M, Barz D, Rupp S, Hube B (2010) Candida glabrata tryptophan-based pigment production via the Ehrlich pathway. Mol Microbiol 76: 25-47.
  • Bulfer SL, Brunzelle JS, Trievel RC (2013) Crystal structure of Saccharomyces cerevisiae Aro8, a putative α-aminoadipate aminotransferase. Protein Sci 22: 1417-1424.
  • Ghosh S, Kebaara BW, Atkin AL, Nickerson KW (2008) Regulation of aromatic alcohol production in Candida albicans. Appl Environ Microbiol 74: 7211-7218.
  • Han Q, Cai T, Tagle D, Li J (2010) Thermal stability, pH dependence and inhibition of four murine kynurenine aminotransferases. BMC Biochem 11: 19-29.
  • Han Q, Cai T, Tagle D, Robinson H, Li J (2009) Substrate specificity and structure of human aminoadipate aminotransferase/kynureine aminotransferase II. Biosci Rep 28: 205-215.
  • Iraqui I, Vissers S, Cartiaux M, Urrestarazu A (1998) Characterisation of Saccharomyces cerevisiae ARO8 and ARO9 genes encoding aromatic aminotransferases I and II reveals a new aminotransferase subfamily. Mol Gen Genet 257: 238-248.
  • Karsten WE, Reyes ZL, Bobyk KD, Cook PF, Chooback L (2011) Mechanism of the aromatic aminotransferase encoded by the Aro8 gene from Saccharomyces cerevisiae. Arch Biochem Biophys 516: 67-74.
  • King RD, Rowland J, Oliver SG, Young M, Aubrey W, Byrne E, Liakata A, Markham MM, Pir P, Soldatova LN, Sparkes A, Whelan KE, Clare A (2009) The automation of science. Science 324: 85-89.
  • Kradolfer P, Niederberger P, Hütter R (1982) Tryptophan degradation in Saccharomyces cerevisiae: characterization of two aromatic aminotransferases. Arch Microbiol 133: 242-248.
  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680-685.
  • Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA, McWilliam H, Valentin F, Wallace IM, Wilm A, Lopez R, Thompson JD, Gibson TJ, Higgins DG (2007) Clustal W and Clustal X version 2.0. Bioinformatics 23: 2947-2948.
  • Matsuda M, Ogur M (1969) Enzymatic and physiological properties of the yeast glutamate-α-ketoadipate transaminase. J Biol Chem 244: 5153-5158.
  • Miyazaki T, Miyazaki J, Yamane H, Nishiyama M (2004) α-Aminoadipate aminotransferase from an extremely thermophilic bacterium, Thermus thermophilus. Microbiology 150: 2327-2334.
  • Okamoto A, Nakai Y, Hayashi H, Hirotsu KK (1998) Crystal Structures of Paracoccus denitrificans aromatic amino acid aminotransferase: a substrate recognition site constructed by rearrangement of hydrogen bond network. J Mol Biol 280: 443-461.
  • Ouchi T, Tomita T, Miyagawa T, Kuzuyama T, Nishiyama M (2009) Dual roles of a conserved pair, Arg23 and Ser20, in recognition of multiple substrates in α-aminoadipate aminotransferase from Thermus thermophilus. Biochem Biophys Res Commun 388: 21-27.
  • Robert X, Gouet P (2014) Deciphering key features in protein structures with the new ENDscript server. Nucleic Acids Res 42: W320-W324.
  • Rossi F, Garavaglia S, Montalbano V, Walsh M, Rizzi M (2008) Crystal structure of human kynurenine aminotransferase II, a drug target for the treatment of schizophrenia. J Biol Chem 283: 3559-3566.
  • Smith DL, Almo SC, Toney MD, Ringe D (1989) 2.8 angstrom resolution crystal structure of an active-site mutant of aspartate aminotransferase from Escherichia coli. Biochemistry 28: 8161-8167.
  • Sung MH, Tanizawa K, Tanaka H, Kuramitsu S, Kagamiyama H, Soda K (1990) Purification and characterization of thermostable aspartate aminotransferase from a thermophilic Bacillus species. J Bacteriol 172: 1345-1351.
  • Tomita T, Miyagawa T, Miyazaki T, Fushinobu S, Kuzuyama T, Nishiyama M (2009) Mechanism for multiple-substrates recognition of a-aminoadipate aminotransferase from Thermus thermophilus. Proteins 75: 348-359.
  • Umemura I, Yanagiya K, Komatsubara S, Sato T, Tosa T (1994) Purification and some properties of alanine aminotransferase from Candida maltosa. Biosci Biotechnol Biochem 58: 283-287.
  • Urrestarazu A, Vissers S, Iraqui I, Grenson M (1998) Phenylalanine- and tyrosine-auxotrophic mutants of Saccharomyces cerevisiae impaired in transamination. Mol Gen Genet 257: 230-237.
  • Ward DE, Kengen SW, van Der Oost J, de Vos WM (2000) Purification and characterization of the alanine aminotransferase from the hyperthermophilic Archaeon Pyrococcus furiosus and its role in alanine production J Bacteriol 182: 2559-2566.
  • Xu H, Andi B, Qian J, West AH, Cook PF (2006) The α-aminoadipate pathway for lysine biosynthesis in fungi. Cell Biochem Biophys 46: 43-64.
Document Type
Publication order reference
Identifiers
YADDA identifier
bwmeta1.element.bwnjournal-article-abpv62p903kz
JavaScript is turned off in your web browser. Turn it on to take full advantage of this site, then refresh the page.