Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Novel polysaccharide–protein-based amphipathic formulations

Abstract

Previous results showed that the cell-surface esterase from Acinetobacter venetianus RAG-1 enhances the emulsification properties of the polymeric bioemulsifier emulsan and its deproteinated derivative apoemulsan (Bach H, Berdichevsky Y, Gutnick D (2003) An exocellular protein from the oil-degrading microbe Acinetobacter venetianus RAG-1 enhances the emulsifying activity of the polymeric bioemulsifier emulsan. Appl Environ Microbiol 69:2608–2615). Here we show that in the presence of the his-tagged recombinant esterase from RAG-1, 18 different polysaccharides from microbial, plant, insect and synthetic sources formed hexadecane-in-water emulsions. Emulsifying activities were distributed over a 13-fold range from over 4800 U/mg protein/mg polysaccharide in the case of apoemulsan to 370 U/mg protein/mg polysaccharide in the case of alginic acid. The stability of the emulsions ranged between 95 and 58%. Emulsions formed in the presence of seven of the polysaccharides exhibited stabilities of over 80%. The esterase from A. calcoaceticus BD4, which shows sequence homology to the RAG-1 esterase, was inactive in emulsification enhancement. The sequence of the RAG-1 esterase was shown to contain two conserved peptide sequences previously shown to be implicated in carbohydrate/polysaccharide binding. A hypothetical model illustrating a possible mode of interaction between the esterase, the apoemulsan and the oil droplet is presented. The complex is presumed to generate a series of “coated” oil droplets which are restricted in their ability to coalesce resulting in a relatively stable emulsion.

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

Fig. 1

References

  1. Bach H, Gutnick DL (2004) Potential applications of bioemulsifiers in the oil industry. In: Vazquez-Duhalt R, Quintero-Ramírez R (eds) Petroleum biotechnology, developments and perspectives. Elsevier BV, Amsterdam, pp 233–281

  2. Bach H, Berdichevsky Y, Gutnick D (2003) An exocellular protein from the oil-degrading microbe Acinetobacter venetianus RAG-1 enhances the emulsifying activity of the polymeric bioemulsifier emulsan. Appl Environ Microbiol 69:2608–2615

  3. Belsky I, Gutnick DL, Rosenberg E (1979) Emulsifier of Arthrobacter RAG-1: determination of emulsifier-bound fatty acids. FEBS Lett 101:175–178

  4. Brzozowski AM, Lawson DM, Turkenburg JP, Bisgaard-Frantzen H, Svendsen A, Borchert TV, Dauter Z, Wilson KS, Davies GJ (2000) Structural analysis of a chimeric bacterial alpha-amylase. High-resolution analysis of native and ligand complexes. Biochemistry 39:9099–9107

  5. Cirigliano MC, Carman GM (1985) Purification and characterization of Liposan, a bioemulsifier from Candida lipolytica. Appl Environ Microbiol 50:846–850

  6. Desai JD, Banat IM (1997) Microbial production of surfactants and their commercial potential. Microbiol Mol Biol Rev 61:47–64

  7. Flint J, Nurizzo D, Harding SE, Longman E, Davies GJ, Gilbert HJ, Bolam DN (2004) Ligand-mediated dimerization of a carbohydrate-binding molecule reveals a novel mechanism for protein–carbohydrate recognition. J Mol Biol 337:417–426

  8. Fromm JR, Hileman RE, Caldwell EE, Weiler JM, Linhardt RJ (1997) Pattern and spacing of basic amino acids in heparin binding sites. Arch Biochem Biophys 343:92–100

  9. Garti N, Reichman D (1994) Surface properties and emulsification activity of galactomannans. Food Hydrocoll 8:155–173

  10. Garti N, Slavin Y, Aserin A (1998) Surface and emulsification properties of a new gum extract from Portulaca oleracea L. Food Hydrocoll 13:145–155

  11. Gutnick DL (1987) The emulsan polymer: perspectives on a microbial capsule as an industrial product. Biopolymers 26:S223–S240

  12. Gutnick DL, Minas W (1987) Perspectives on microbial surfactants. Biochem Soc Trans 15 Suppl:22S–35S

  13. Imamura H, Fushinobu S, Yamamoto M, Kumasaka T, Jeon BS, Wakagi T, Matsuzawa H (2003) Crystal structures of 4-alpha-glucanotransferase from Thermococcus litoralis and its complex with an inhibitor. J Biol Chem 278:19378–19386

  14. Juni EJ, Janik A (1969) Transformation of Acinetobacter calcoaceticum (Bacterium anitatum). J Bacteriol 98:281–288

  15. Kanai R, Haga K, Akiba T, Yamane K, Harata K (2004) Role of Phe283 in enzymatic reaction of cyclodextrin glycosyltransferase from alkalophilic Bacillus sp.1011: substrate binding and arrangement of the catalytic site. Protein Sci 13:457–465

  16. Kaplan N, Zosim Z, Rosenberg E (1987) Reconstitution of emulsifying activity of Acinetobacter calcoaceticus BD4 emulsan by using pure polysaccharide and protein. Appl Environ Microbiol 53:440–446

  17. Kok RG, Christoffels VM, Vosman B, Hellingwerf KJ (1993) Growth-phase-dependent expression of the lipolytic system of Acinetobacter calcoaceticus BD413: cloning of a gene encoding one of the esterases. J Gen Microbiol 139:2329–2342

  18. Ophir T, Gutnick DL (1994) A role for exopolysaccharides in the protection of microorganisms from desiccation. Appl Environ Microbiol 60:740–745

  19. Papanikolau Y, Prag G, Tavlas G, Vorgias CE, Oppenheim AB, Petratos K (2001) High resolution structural analyses of mutant chitinase A complexes with substrates provide new insight into the mechanism of catalysis. Biochemistry 40:11338–11343

  20. Randall RC, Philips GO, Williams PA (1988) The role of the proteinaceous component on the emulsifying properties of gum arabic. Food Hydrocoll 2:131–140

  21. Reisfeld A, Rosenberg E, Gutnick DL (1972) Microbial degradation of crude oil: factors affecting the dispersion in sea water by mixed and pure cultures. Appl Microbiol 24:363–368

  22. Rosenberg E, Zuckerberg A, Rubinovitz C, Gutnick DL (1979) Emulsifier of Arthrobacter RAG-1: isolation and emulsifying properties. Appl Environ Microbiol 37:402–408

  23. Sanz-Aparicio J, Hermoso JA, Martinez-Ripoll M, Lequerica JL, Polaina J (1998) Crystal structure of beta-glucosidase A from Bacillus polymyxa: insights into the catalytic activity in family 1 glycosyl hydrolases. J Mol Biol 275:491–502

  24. Shabtai Y, Gutnick DL (1985) Exocellular esterase and emulsan release from the cell surface of Acinetobacter calcoaceticus. J Bacteriol 161:1176–1181

  25. Shoham Y, Rosenberg M, Rosenberg E (1983) Bacterial degradation of emulsan. Appl Environ Microbiol 46:573–579

  26. Stout V, Torres-Cabassa A, Maurizi MR, Gutnick D, Gottesman S (1991) RcsA, an unstable positive regulator of capsular polysaccharide synthesis. J Bacteriol 173:1738–1747

  27. Vyas NK (1991) Atomic features of protein–carbohydrate interactions. Curr Opin Struct Biol 1:732–740

Download references

Acknowledgements

This work was supported in part by a grant from the CDR project of the US A.I.D. We thank Rina Avigad for excellent technical assistance. Thanks also to David Nakar for ideas and useful suggestions. HB was a predoctoral fellow in the Department of Molecular Microbiology and Biotechnology at Tel-Aviv University.

Author information

Correspondence to David L. Gutnick.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Bach, H., Gutnick, D.L. Novel polysaccharide–protein-based amphipathic formulations. Appl Microbiol Biotechnol 71, 34–38 (2006). https://doi.org/10.1007/s00253-005-0149-9

Download citation

Keywords

  • Polysaccharide
  • Alginic Acid
  • Emulsan
  • Hexadecane
  • Acinetobacter