Abstract
Escherichia coli K4 synthesizes a capsular polysaccharide (CPS) consisting of a fructose-branched chondroitin (GalNAc-GlcA(fructose)n), which is a biosynthetic precursor of chondroitin sulfate. Here, the role of kfoE in the modification of the chondroitin backbone was investigated using knock-out and recombinant complementation experiments. kfoE disruption and complementation had no significant effect on cell growth. CPS production was increased by 15 % in the knock-out strain, and decreased by 21 % in the knock-out strain complemented with recombinant kfoE. CPS extracted from the knock-out strain was chondroitin, whereas CPS extracted from the complemented strain was a fructose-branched chondroitin. The results demonstrated that the kfoE gene product altered the fructose group at the C3 position of the GlcA residue during production of K4CPS.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Bahrani-Mougeot FK, Buckles EL, Lockatell CV, Hebel JR, Johnson DE, Tang CM, Donnenberg MS (2002) Type 1 fimbriae and extracellular polysaccharides are preeminent uropathogenic Escherichia coli virulence determinants in the murine urinary tract. Mol Microbiol 45:1079–1093
Bedini E, De Castro C, De Rosa M, Di Nola A, Iadonisi A, Restaino OF, Schiraldi C, Parrilli M (2011) A microbiological-chemical strategy to produce chondroitin sulfate A, C. Angew Chem Int Ed Engl 50:6160–6163
Bedini E, De Castro C, De Rosa M, Di Nola A, Restaino OF, Schiraldi C, Parrilli M (2012) Semi-synthesis of unusual chondroitin sulfate polysaccharides containing GlcA(3-O-sulfate) or GlcA(2,3-di-O-sulfate) units. Chemistry 18:2123–2130
Bitter T, Muir HM (1962) A modified uronic acid carbazole reaction. Anal Biochem 4:330–334
Cimini D, Restaino OF, Catapano A, De Rosa M, Schiraldi C (2010) Production of capsular polysaccharide from Escherichia coli K4 for biotechnological applications. Appl Microbiol Biotechnol 85:1779–1787
Cimini D, De Rosa M, Carlino E, Ruggiero A, Schiraldi C (2013) Homologous overexpression of RfaH in E. coli K4 improves the production of chondroitin-like capsular polysaccharide. Microb Cell Fact 12:46
Datsenko KA, Wanner BL (2000) One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci USA 97:6640–6645
Hannesson HH, HagnerMcWhirter A, Tiedemann K, Lindahl U, Malmstrom A (1996) Biosynthesis of dermatan sulphate. Defructosylated Escherichia coli K4 capsular polysaccharide as a substrate for the d-glucuronyl C-5 epimerase, and an indication of a two-base reaction mechanism. Biochem J 313:589–596
Hodson N, Griffiths G, Cook N, Pourhossein M, Gottfridson E, Lind T, Lidholt K, Roberts IS (2000) Identification that KfiA, a protein essential for the biosynthesis of the Escherichia coli K5 capsular polysaccharide, is an alpha-UDP-GlcNAc glycosyltransferase. The formation of a membrane-associated K5 biosynthetic complex requires KfiA, KfiB, and KfiC. J Biol Chem 275:27311–27315
Iovu M, Dumais G, du Souich P (2008) Anti-inflammatory activity of chondroitin sulfate. Osteoarthr Cartilage 16(Suppl 3):S14–S18
Karst NA, Linhardt RJ (2003) Recent chemical and enzymatic approaches to the synthesis of glycosaminoglycan oligosaccharides. Curr Med Chem 10:1993–2031
Krahulec J, Krahulcova J, Medova M, Velebny V (2005) Influence of KfoG on capsular polysaccharide structure in Escherichia coli K4 strain. Mol Biotechnol 30:129–134
Kubo M, Ando K, Mimura T, Matsusue Y, Mori K (2009) Chondroitin sulfate for the treatment of hip and knee osteoarthritis: current status and future trends. Life Sci 85:477–483
Lidholt K, Fjelstad M (1997) Biosynthesis of the Escherichia coli K4 capsule polysaccharide. A parallel system for studies of glycosyltransferases in chondroitin formation. J Biol Chem 272:2682–2687
Lubineau A, Bonnaffe D (1999) Access to molecular diversity in glycosaminoglycans: combinatorial synthesis of eight chondroitin sulfate disaccharides. Eur J Org Chem 1999:2523–2532
Manzoni M, Bergomi S, Molinari F, Cavazzoni V (1996) Production and purification of an extracellularly produced K4 polysacchariide from Escherichia coli. Biotechnol Lett 18:383–386
Ninomiya T, Sugiura N, Tawada A, Sugimoto K, Watanabe H, Kimata K (2002) Molecular cloning and characterization of chondroitin polymerase from Escherichia coli strain K4. J Biol Chem 277:21567–21575
Osawa T, Sugiura N, Shimada H, Hirooka R, Tsuji A, Shirakawa T, Fukuyama K, Kimura M, Kimata K, Kakuta Y (2009) Crystal structure of chondroitin polymerase from Escherichia coli K4. Biochem Biophys Res Commun 378:10–14
Petit C, Rigg GP, Pazzani C, Smith A, Sieberth V, Stevens M, Boulnois G, Jann K, Roberts IS (1995) Region 2 of the Escherichia coli K5 capsule gene cluster encoding proteins for the biosynthesis of the K5 polysaccharide. Mol Microbiol 17:611–620
Rodriguez ML, Jann B, Jann K (1988) Structure and serological characteristics of the capsular K4 antigen of Escherichia coli O5:k4:H4, a fructose-containing polysaccharide with a chondroitin backbone. Eur J Biochem 177:117–124
Ronca F, Palmieri L, Panicucci P, Ronca G (1998) Anti-inflammatory activity of chondroitin sulfate. Osteoarthr Cartil 6(Suppl A):14–21
Snyder JA, Haugen BJ, Buckles EL, Lockatell CV, Johnson DE, Donnenberg MS, Welch RA, Mobley HL (2004) Transcriptome of uropathogenic Escherichia coli during urinary tract infection. Infect Immun 72:6373–6381
Sobhany M, Kakuta Y, Sugiura N, Kimata K, Negishi M (2008) The chondroitin polymerase K4CP and the molecular mechanism of selective bindings of donor substrates to two active sites. J Biol Chem 283:32328–32333
Whitfield C (2006) Biosynthesis and assembly of capsular polysaccharides in Escherichia coli. Annu Rev Biochem 75:39–68
Wu Q, Yang A, Zou W, Duan Z, Liu J, Chen J, Liu L (2013) Transcriptional engineering of Escherichia coli K4 for fructosylated chondroitin production. Biotechnol Prog 29:1140–1149
Yu H, Stephanopoulos G (2008) Metabolic engineering of Escherichia coli for biosynthesis of hyaluronic acid. Metab Eng 10:24–32
Zhang C, Liu L, Teng L, Chen J, Liu J, Li J, Du G, Chen J (2012) Metabolic engineering of Escherichia coli BL21 for biosynthesis of heparosan, a bioengineered heparin precursor. Metab Eng 14:521–527
Acknowledgments
This work was financially supported by Program for the Key Technologies R & D Program of Jiangsu Province (BE2012617), the Program for Young Talents in China, the provincal outstanding youth foundation of Jiangsu Province (BK2012002).
Author information
Authors and Affiliations
Corresponding author
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Liu, J., Yang, A., Liu, J. et al. KfoE encodes a fructosyltransferase involved in capsular polysaccharide biosynthesis in Escherichia coli K4. Biotechnol Lett 36, 1469–1477 (2014). https://doi.org/10.1007/s10529-014-1502-9
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10529-014-1502-9