Skip to main content
SpringerLink
Log in

Genomic data mining reveals the transaminase repertoire of Komagataella phaffii (Pichia pastoris) strain GS115 and supports a systematic nomenclature

  • Research Article
  • Published:
Journal of Genetics Aims and scope Submit manuscript

Abstract

Transaminases are an industrially important class of enzyme, due to their ability to catalyse amination reactions for production of chiral amines, and are key building blocks of small molecule pharmaceuticals. We analysed the genome of strain GS115 of the methylotrophic yeast Komagataella phaffii, formerly known as Pichia pastoris, to identify the transaminase genes and propose a systematic nomenclature based on both phylogeny and structuro-functional features. K. phaffii is an increasingly attractive industrial host cell due to its ability to grow to high biomass, up to 60% wet cell weight by volume, using methanol as carbon source and inducer of transgene expression. Thirty-nine UniProt database hits were reduced to 19 on the basis of sequence similarity and hidden Markov model. Of the 19 genes, the open-reading frames of three (KpTam I-II.1b, KpTam I-II.7 and KpTam V.2) had strong homology with no characterized protein and four (KpTam III.1a, KpTam III.1b, KpTam III.2a and KpTam III.2b) had relatively high sequence similarity to ω-type transaminases, a subtype that typically accepts the broadest range of substrates. Comparison with Saccharomyces cerevisiae S288C suggested functions for KpTam I-II.1b and KpTam I-II.7. K. phaffii GS115 was originally generated by mutagenesis of K. phaffii CBS7435 and comparison revealed that one transaminase gene may have been deleted during this mutagenesis. These insights can advance fundamental understanding of yeast biology and can inform industrial screening and engineering of yeast transaminases.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Figure 1
Figure 2

Similar content being viewed by others

References

  • Bairoch A. 2000 The ENZYME database in 2000. Nucleic Acids Res. 28, 304–305.

    Article  CAS  Google Scholar 

  • Bollok M., Resina D., Valero F. and Ferrer P. 2009 Recent patents on the Pichia pastoris expression system: expanding the toolbox for recombinant protein production. Recent Pat. Biotechnol. 3,192–201

    Article  CAS  Google Scholar 

  • Cho B. K., Cho H. J., Park S. H., Yun H. and Kim B. G. 2003 Simultaneous synthesis of enantiomerically pure (S)-amino acids and (R)-amines using coupled transaminase reactions. Biotechnol. Bioeng. 81, 783–789.

    Article  CAS  Google Scholar 

  • De Schutter K., Lin Y. C., Tiels P., Van Hecke A., Glinka S., Weber-Lehmann J. et al. 2009 Genome sequence of the recombinant protein production host Pichia pastoris. Nat. Biotechnol. 27, 561–566.

    Article  Google Scholar 

  • Engel S. R., Dietrich F. S., Fisk D. G., Binkley G., Balakrishnan R., Costanzo M. C. et al. 2014 The reference genome sequence of Saccharomyces cerevisiae: then and now. G3 (Bethesda) 4, 389–398.

    Article  Google Scholar 

  • Finn R. D., Clements J., Arndt W., Miller B. L., Wheeler T. J., Schreiber F. et. al. 2015 HMMER web server: 2015 update. Nucleic Acids Res. 43, W30–W38.

    Article  CAS  Google Scholar 

  • Finn R. D., Coggill P., Eberhardt R. Y., Eddy S. R., Mistry J., Mitchell A. L. et al. 2016 The Pfam protein families database: towards a more sustainable future. Nucleic Acids Res. 44, D279–D285.

    Article  CAS  Google Scholar 

  • Goffeau A., Barrell B. G., Bussey H., Davis R. W., Dujon B., Feldmann H. et al. 1996 Life with 6000 genes. Science 274, 563–567.

    Article  Google Scholar 

  • Henikoff S. and Henikoff J. G. 1992 Amino acid substitution matrices from protein blocks. Proc. Natl. Acad. Sci. USA 89, 10915–10919.

    Article  CAS  Google Scholar 

  • Hohne M., Schatzle S., Jochens H., Robins K. and Bornscheuer U. T. 2010 Rational assignment of key motifs for function guides in silico enzyme identification. Nat. Chem. Biol. 6, 807–813.

    Article  Google Scholar 

  • Huson D. H. and Scornavacca C. 2012 Dendroscope 3: an interactive tool for rooted phylogenetic trees and networks. Syst. Biol. 61, 1061–1067.

    Article  Google Scholar 

  • Hwang B-Y., Cho B-K., Yun H., Koteshwar K. and Kim B.-G. 2005 Revisit of aminotransferase in the genomic era and its application to biocatalysis. J. Mol. Cat. B: Enzym. 37, 47–55.

    Article  CAS  Google Scholar 

  • Ingram C. U., Bommer M., Smith M. E., Dalby P. A., Ward J. M., Hailes H. et al. 2007 One-pot synthesis of amino-alcohols using a de-novo transketolase and beta-alanine: pyruvate transaminase pathway in Escherichia coli. Biotechnol. Bioeng. 96, 559–569.

    Article  CAS  Google Scholar 

  • Invitrogen 2010 Multi-Copy Pichia Expression kit. For the isolation and expression of recombinant proteins from Pichia pastoris strains containing multiple copies of a particular gene. User Manual part no. 25–0170.

  • Kaulmann U., Smithies K., Smith M. E. B., Hailes H. C. and Ward J. M. 2007 Substrate spectrum of ω-transaminase from Chromobacterium violaceum DSM30191 and its potential for biocatalysis. Enzyme Microb. Technol. 41, 628–637.

    Article  CAS  Google Scholar 

  • Krogh A., Brown M., Mian I. S., Sjolander K. and Haussler D 1994 Hidden Markov models in computational biology. Applications to protein modeling. J. Mol. Biol. 235, 1501–1531.

    Article  CAS  Google Scholar 

  • Küberl A., Schneider J., Thallinger G. G., Anderl I., Wibberg D., Hajek T. et al. 2011 High-quality genome sequence of Pichia pastoris CBS7435. J. Biotechnol. 154, 312–320.

    Article  Google Scholar 

  • Kurtzman C. P. 2005 Description of Komagataella phaffii sp. nov. and the transfer of Pichia pseudopastoris to the methylotrophic yeast genus Komagataella. Int. J. Syst. Evol. Microbiol. 55, 973–976.

    Article  CAS  Google Scholar 

  • Larkin M. A, Blackshields G, Brown N. P., Chenna R., McGettigan P. A. et al. 2007 Clustal W and Clustal X version 2.0. Bioinformatics 23, 2947–2948.

    Article  CAS  Google Scholar 

  • Macauley-Patrick S., Fazenda M. L., McNeil B. and Harvey L. M. 2005 Heterologous protein production using the Pichia pastoris expression system. Yeast 22, 249–270.

    Article  CAS  Google Scholar 

  • Malik M. S., Park E. S. and Shin J. S. 2012 Features and technical applications of omega-transaminases. Appl. Microbiol. Biotechnol. 94, 1163–1171.

    Article  CAS  Google Scholar 

  • Mattanovich D., Callewaert N., Rouze P., Lin Y. C., Graf A., Redl A. et. al 2009 Open access to sequence: browsing the Pichia pastoris genome. Microb. Cell Factories 8, 53.

    Article  Google Scholar 

  • Mehta P. K., Hale T. I. and Christen P. 1993 Aminotransferases: demonstration of homology and division into evolutionary subgroups. Eur. J. Biochem. 214, 549–561.

    Article  CAS  Google Scholar 

  • Qian B. and Goldstein R. A. 2003 Detecting distant homologs using phylogenetic tree-based HMMs. Proteins 52, 446–453.

    Article  CAS  Google Scholar 

  • Richardson S. M., Mitchell L. A., Stracquadanio G., Yang K., Dymond J. S., DiCarlo J. E. et al. 2017 Design of a synthetic yeast genome. Science 355, 1040–1044.

    Article  CAS  Google Scholar 

  • Robert X. and Gouet P. 2014 Deciphering key features in protein structures with the new ENDscript server. Nucleic Acids Res. 42, W320–W324.

    Article  CAS  Google Scholar 

  • Sayer C., Isupov M. N. and Littlechild J. A. 2007 Crystallization and preliminary X-ray diffraction analysis of omega-amino acid: puruvate transaminase from Chromobacterium violaceum. Acta Crystallogr. Sect. F Strct. Biol. Cryst. Commun. 63, 117–119.

    Article  CAS  Google Scholar 

  • Shen B. W., Hennig M., Hohenester E., Jansonius J. N. and Schirmer T. 1998 Crystal structure of human recombinant ornithine aminotransferase. J. Mol. Biol. 277, 81–102.

    Article  CAS  Google Scholar 

  • Shin J. S., Yun H., Jang J. W, Park I. and Kim B. G. 2003 Purification, characterization, and molecular cloning of a novel amine:pyruvate transaminase from Vibrio fluvialis JS17. Appl. Microbiol. Biotechnol. 61, 463–471.

    Article  CAS  Google Scholar 

  • Templar A., Woodhouse S., Keshavarz-Moore E. and Nesbeth D. N. 2016 Influence of Pichia pastoris cellular material on polymerase chain reaction performance as a synthetic biology standard for genome monitoring. J. Microbiol. Methods 127, 111–122.

    Article  CAS  Google Scholar 

  • Valli M., Tatto N. E., Peymann A., Gruber C., Landes N., Ekker H. et al. 2016 Curation of the genome annotation of Pichia pastoris (Komagataella phaffii) CBS7435 from gene level to protein function. FEMS Yeast Res. 16, fow051.

    Article  Google Scholar 

  • Van der Klei I. J., Yurimoto H., Sakai Y. and Veenhuis M. 2006 The significance of peroxisomes in methanol metabolism in methylotrophic yeast. Biochim. Biophys. Acta 1763, 1453–1462.

    Article  Google Scholar 

  • Ward J. and Wohlgemuth R. 2010 High-yield biocatalytic amination reactions in organic synthesis. Curr. Org. Chem. 14, 1914–1927.

    Article  CAS  Google Scholar 

  • Waterhouse A. M., Procter J. B., Martin D. M., Clamp M. and Barton G. J. 2009 Jalview Version 2–a multiple sequence alignment editor and analysis workbench. Bioinformatics 25, 1189–1191.

    Article  CAS  Google Scholar 

  • Wei Y. C., Braun-Galleani S., Henriquez M. J., Bandara S. and Nesbeth D. N. 2018 Biotransformation of beta-Hydroxypyruvate and Glycolaldehyde to L-Erythrulose by Pichia pastoris strain GS115 overexpressing native Transketolase. Biotechnol. Prog. 34, 99–106.

    Article  CAS  Google Scholar 

  • Yonaha K., Nishie M. and Aibara S. 1992 The primary structure of omega-amino acid:pyruvate aminotransferase. J. Biol. Chem. 267, 12506–12510.

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by the Government of Chile and its Program CONICYT-Becas Chile (72120390), who supported the doctoral studies of M.-J.H., and the ERA-IB IPCRES Consortium, funded in the UK by the BBSRC (BB/M004880/1), who supported all other authors.

Author information

Authors and Affiliations

  1. Department of Biochemical Engineering, University College London, Bernard Katz Building, London, WC1E 6BT, UK

    Maria-José Henríquez, Rosalía Paula Cardós-Elena & Darren Nicholas Nesbeth

Authors
  1. Maria-José Henríquez
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Rosalía Paula Cardós-Elena
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Darren Nicholas Nesbeth
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Darren Nicholas Nesbeth.

Additional information

Corresponding editor: H. A. Ranganath

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Henríquez, MJ., Cardós-Elena, R.P. & Nesbeth, D.N. Genomic data mining reveals the transaminase repertoire of Komagataella phaffii (Pichia pastoris) strain GS115 and supports a systematic nomenclature. J Genet 99, 49 (2020). https://doi.org/10.1007/s12041-020-01201-1

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1007/s12041-020-01201-1

Keywords

Search

Navigation