A minimal peptide substrate in biotin holoenzyme synthetase-catalyzed biotinylation (original) (raw)

Abstract

The Escherichia coli biotin holoenzyme synthetase, BirA, catalyzes transfer of biotin to the epsilon amino group of a specific lysine residue of the biotin carboxyl carrier protein (BCCP) subunit of acetyl-CoA carboxylase. Sequences of naturally biotinylated substrates are highly conserved across evolutionary boundaries, and cross-species biotinylation has been demonstrated in several systems. To define the minimal substrate requirements in BirA-catalyzed biotinylation, we have measured the kinetics of modification of a 23-residue peptide previously identified by combinatorial methods. Although the sequence of the peptide bears little resemblance to the biotinylated sequence in BCCP, it is enzymatically biotinylated in vivo. Rates of biotin transfer to the 23-residue peptide are similar to those determined for BCCP. To further elucidate the sequence requirements for biotinylation, transient kinetic measurements were performed on a series of amino- and carboxy-terminal truncations of the 23-mer. The results, determined by stopped-flow fluorescence, allowed identification of a 14-residue peptide as the minimum required sequence. Additional support was obtained using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometric analysis of peptides that had been incubated with an excess of biotinyl-5'-adenylate intermediate and catalytic amounts of BirA. Results of these measurements indicate that while kinetically inactive truncations showed no significant shift in molecular mass to the values expected for biotinylated species, kinetically active truncations exhibited 100% biotinylation. The specificity constant (k(cat)/Km) governing BirA-catalyzed biotinylation of the 14-mer minimal substrate is similar to that determined for the natural substrate, BCCP. We conclude that the 14-mer peptide efficiently mimics the biotin acceptor function of the much larger protein domain normally recognized by BirA.

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Selected References

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  1. Abbott J., Beckett D. Cooperative binding of the Escherichia coli repressor of biotin biosynthesis to the biotin operator sequence. Biochemistry. 1993 Sep 21;32(37):9649–9656. doi: 10.1021/bi00088a017. [DOI] [PubMed] [Google Scholar]
  2. Altman J. D., Moss P. A., Goulder P. J., Barouch D. H., McHeyzer-Williams M. G., Bell J. I., McMichael A. J., Davis M. M. Phenotypic analysis of antigen-specific T lymphocytes. Science. 1996 Oct 4;274(5284):94–96. doi: 10.1126/science.274.5284.94. [DOI] [PubMed] [Google Scholar]
  3. Athappilly F. K., Hendrickson W. A. Structure of the biotinyl domain of acetyl-coenzyme A carboxylase determined by MAD phasing. Structure. 1995 Dec 15;3(12):1407–1419. doi: 10.1016/s0969-2126(01)00277-5. [DOI] [PubMed] [Google Scholar]
  4. Brocklehurst S. M., Perham R. N. Prediction of the three-dimensional structures of the biotinylated domain from yeast pyruvate carboxylase and of the lipoylated H-protein from the pea leaf glycine cleavage system: a new automated method for the prediction of protein tertiary structure. Protein Sci. 1993 Apr;2(4):626–639. doi: 10.1002/pro.5560020413. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chandler C. S., Ballard F. J. Regulation of the breakdown rates of biotin-containing proteins in Swiss 3T3-L1 cells. Biochem J. 1988 May 1;251(3):749–755. doi: 10.1042/bj2510749. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cronan J. E., Jr Biotination of proteins in vivo. A post-translational modification to label, purify, and study proteins. J Biol Chem. 1990 Jun 25;265(18):10327–10333. [PubMed] [Google Scholar]
  7. Cull M. G., Miller J. F., Schatz P. J. Screening for receptor ligands using large libraries of peptides linked to the C terminus of the lac repressor. Proc Natl Acad Sci U S A. 1992 Mar 1;89(5):1865–1869. doi: 10.1073/pnas.89.5.1865. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Duplay P., Bedouelle H., Fowler A., Zabin I., Saurin W., Hofnung M. Sequences of the malE gene and of its product, the maltose-binding protein of Escherichia coli K12. J Biol Chem. 1984 Aug 25;259(16):10606–10613. [PubMed] [Google Scholar]
  9. Edelhoch H. Spectroscopic determination of tryptophan and tyrosine in proteins. Biochemistry. 1967 Jul;6(7):1948–1954. doi: 10.1021/bi00859a010. [DOI] [PubMed] [Google Scholar]
  10. Fall R. R. Analysis of microbial biotin proteins. Methods Enzymol. 1979;62:390–398. doi: 10.1016/0076-6879(79)62246-2. [DOI] [PubMed] [Google Scholar]
  11. Kim D. R., McHenry C. S. Biotin tagging deletion analysis of domain limits involved in protein-macromolecular interactions. Mapping the tau binding domain of the DNA polymerase III alpha subunit. J Biol Chem. 1996 Aug 23;271(34):20690–20698. doi: 10.1074/jbc.271.34.20690. [DOI] [PubMed] [Google Scholar]
  12. Knowles J. R. The mechanism of biotin-dependent enzymes. Annu Rev Biochem. 1989;58:195–221. doi: 10.1146/annurev.bi.58.070189.001211. [DOI] [PubMed] [Google Scholar]
  13. LANE M. D., ROMINGER K. L., YOUNG D. L., LYNEN F. THE ENZYMATIC SYNTHESIS OF HOLOTRANSCARBOXYLASE FROM APOTRANSCARBOXYLASE AND (+)-BIOTIN. II. INVESTIGATION OF THE REACTION MECHANISM. J Biol Chem. 1964 Sep;239:2865–2871. [PubMed] [Google Scholar]
  14. Leon-Del-Rio A., Gravel R. A. Sequence requirements for the biotinylation of carboxyl-terminal fragments of human propionyl-CoA carboxylase alpha subunit expressed in Escherichia coli. J Biol Chem. 1994 Sep 16;269(37):22964–22968. [PubMed] [Google Scholar]
  15. León-Del-Rio A., Leclerc D., Akerman B., Wakamatsu N., Gravel R. A. Isolation of a cDNA encoding human holocarboxylase synthetase by functional complementation of a biotin auxotroph of Escherichia coli. Proc Natl Acad Sci U S A. 1995 May 9;92(10):4626–4630. doi: 10.1073/pnas.92.10.4626. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Murtif V. L., Samols D. Mutagenesis affecting the carboxyl terminus of the biotinyl subunit of transcarboxylase. Effects on biotination. J Biol Chem. 1987 Aug 25;262(24):11813–11816. [PubMed] [Google Scholar]
  17. Nenortas E., Beckett D. Purification and characterization of intact and truncated forms of the Escherichia coli biotin carboxyl carrier subunit of acetyl-CoA carboxylase. J Biol Chem. 1996 Mar 29;271(13):7559–7567. doi: 10.1074/jbc.271.13.7559. [DOI] [PubMed] [Google Scholar]
  18. Palczewski K., Buczyłko J., Kaplan M. W., Polans A. S., Crabb J. W. Mechanism of rhodopsin kinase activation. J Biol Chem. 1991 Jul 15;266(20):12949–12955. [PubMed] [Google Scholar]
  19. Reddy D. V., Rothemund S., Shenoy B. C., Carey P. R., Sönnichsen F. D. Structural characterization of the entire 1.3S subunit of transcarboxylase from Propionibacterium shermanii. Protein Sci. 1998 Oct;7(10):2156–2163. doi: 10.1002/pro.5560071013. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Reiss Y., Stradley S. J., Gierasch L. M., Brown M. S., Goldstein J. L. Sequence requirement for peptide recognition by rat brain p21ras protein farnesyltransferase. Proc Natl Acad Sci U S A. 1991 Feb 1;88(3):732–736. doi: 10.1073/pnas.88.3.732. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Robinson B. H., Oei J., Saunders M., Gravel R. [3H]biotin-labeled proteins in cultured human skin fibroblasts from patients with pyruvate carboxylase deficiency. J Biol Chem. 1983 May 25;258(10):6660–6664. [PubMed] [Google Scholar]
  22. Ross J. B., Senear D. F., Waxman E., Kombo B. B., Rusinova E., Huang Y. T., Laws W. R., Hasselbacher C. A. Spectral enhancement of proteins: biological incorporation and fluorescence characterization of 5-hydroxytryptophan in bacteriophage lambda cI repressor. Proc Natl Acad Sci U S A. 1992 Dec 15;89(24):12023–12027. doi: 10.1073/pnas.89.24.12023. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Samols D., Thornton C. G., Murtif V. L., Kumar G. K., Haase F. C., Wood H. G. Evolutionary conservation among biotin enzymes. J Biol Chem. 1988 May 15;263(14):6461–6464. [PubMed] [Google Scholar]
  24. Schatz P. J. Use of peptide libraries to map the substrate specificity of a peptide-modifying enzyme: a 13 residue consensus peptide specifies biotinylation in Escherichia coli. Biotechnology (N Y) 1993 Oct;11(10):1138–1143. doi: 10.1038/nbt1093-1138. [DOI] [PubMed] [Google Scholar]
  25. Shenoy B. C., Paranjape S., Murtif V. L., Kumar G. K., Samols D., Wood H. G. Effect of mutations at Met-88 and Met-90 on the biotination of Lys-89 of the apo 1.3S subunit of transcarboxylase. FASEB J. 1988 Jun;2(9):2505–2511. doi: 10.1096/fasebj.2.9.3131174. [DOI] [PubMed] [Google Scholar]
  26. Shenoy B. C., Xie Y., Park V. L., Kumar G. K., Beegen H., Wood H. G., Samols D. The importance of methionine residues for the catalysis of the biotin enzyme, transcarboxylase. Analysis by site-directed mutagenesis. J Biol Chem. 1992 Sep 15;267(26):18407–18412. [PubMed] [Google Scholar]
  27. Smith P. A., Tripp B. C., DiBlasio-Smith E. A., Lu Z., LaVallie E. R., McCoy J. M. A plasmid expression system for quantitative in vivo biotinylation of thioredoxin fusion proteins in Escherichia coli. Nucleic Acids Res. 1998 Mar 15;26(6):1414–1420. doi: 10.1093/nar/26.6.1414. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Songyang Z., Carraway K. L., 3rd, Eck M. J., Harrison S. C., Feldman R. A., Mohammadi M., Schlessinger J., Hubbard S. R., Smith D. P., Eng C. Catalytic specificity of protein-tyrosine kinases is critical for selective signalling. Nature. 1995 Feb 9;373(6514):536–539. doi: 10.1038/373536a0. [DOI] [PubMed] [Google Scholar]
  29. Tatsumi H., Fukuda S., Kikuchi M., Koyama Y. Construction of biotinylated firefly luciferases using biotin acceptor peptides. Anal Biochem. 1996 Dec 1;243(1):176–180. doi: 10.1006/abio.1996.0498. [DOI] [PubMed] [Google Scholar]
  30. Tissot G., Douce R., Alban C. Evidence for multiple forms of biotin holocarboxylase synthetase in pea (Pisum sativum) and in Arabidopsis thaliana: subcellular fractionation studies and isolation of a cDNA clone. Biochem J. 1997 Apr 1;323(Pt 1):179–188. doi: 10.1042/bj3230179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Tsao K. L., DeBarbieri B., Michel H., Waugh D. S. A versatile plasmid expression vector for the production of biotinylated proteins by site-specific, enzymatic modification in Escherichia coli. Gene. 1996 Feb 22;169(1):59–64. doi: 10.1016/0378-1119(95)00762-8. [DOI] [PubMed] [Google Scholar]
  32. Wilson K. P., Shewchuk L. M., Brennan R. G., Otsuka A. J., Matthews B. W. Escherichia coli biotin holoenzyme synthetase/bio repressor crystal structure delineates the biotin- and DNA-binding domains. Proc Natl Acad Sci U S A. 1992 Oct 1;89(19):9257–9261. doi: 10.1073/pnas.89.19.9257. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Yao X., Wei D., Soden C., Jr, Summers M. F., Beckett D. Structure of the carboxy-terminal fragment of the apo-biotin carboxyl carrier subunit of Escherichia coli acetyl-CoA carboxylase. Biochemistry. 1997 Dec 9;36(49):15089–15100. doi: 10.1021/bi971485f. [DOI] [PubMed] [Google Scholar]