Microbial glycosaminoglycan glycosyltransferases

Glycobiology vol. 12 no. 1 pp. 9R–16R, 2002 MINI REVIEW Microbial glycosaminoglycan glycosyltransferases Paul L. DeAngelis Department of Biochemistry and Molecular Biology, Oklahoma Center for Medical Glycobiology, University of Oklahoma Health Sciences Center, 940 Stanton L. Young Blvd., Oklahoma City, OK 73104 Accepted on October 4, 2001 Glycosaminoglycans, a class of linear polysaccharides composed of repeating disaccharide units containing a hexosamine, are important carbohydrates found in many organisms. Vertebrates utilize glycosaminoglycans in structural, recognition, adhesion, and signaling roles. Certain pathogenic bacteria produce extracellular capsules composed of glycosaminoglycans or glycosaminoglycanlike polymers that enhance the microbes’ ability to infect or to colonize the host. In the period from 1993 to 2001, bacterial enzymes were discovered that catalyze the polymerization of the repeating unit of hyaluronan, chondroitin, or N-acetylheparosan (unsulfated, unepimerized heparin). Depending on the specific carbohydrate and the microorganism, either a dual-action enzyme (synthase) that transfers two distinct monosaccharides or a pair of single-action transferases are utilized to synthesize the glycosaminoglycan polymer. Current views on the enzymology, structures, potential evolution, and the roles of the known glycosyltransferases from Streptococcus, Pasteurella, and Escherichia are discussed. Key words: chondroitin/heparin/heparosan/hyaluronan or hyaluronic acid/polysaccharide Bacterial capsules and glycosaminoglycans Pathogenic bacteria are notorious for their ability to surmount host defenses by producing a wide gamut of virulence factors that enhance microbial infectivity and/or persistence. One such factor is the capsule, an extracellular polymer coating surrounding the microbial cell (reviewed in Roberts, 1996). Pathogens that lose the ability to produce a capsule are often attenuated or avirulent. The majority of described capsules are composed of long anionic polysaccharide chains, but neutral polysaccharides as well as proteinaceous components are also observed in some cases. Hundreds of structures have been reported from animal and plant pathogens, but of special interest are capsular polymers chemically identical or similar to host molecules. In particular, both vertebrates and certain microbes produce glycosaminoglycans (GAGs), linear polysaccharides composed of repeating disaccharide units containing a derivative of an amino sugar (either glucosamine or galactosamine). © 2002 Oxford University Press Hyaluronan (HA), chondroitin, and heparan sulfate/heparin contain a uronic acid as the other component of the disaccharide repeat, and keratan contains a galactose (Table I). Vertebrates can contain all four types of GAGs, but the polysaccharide chain is often further modified after sugar polymerization. One or more modifications, including O-sulfation of certain hydroxyls, deacetylation and subsequent N-sulfation, or epimerization of glucuronic acid to iduronic acid, are found in most GAGs except HA. An amazing variety of distinct structures have been reported for chondroitin sulfate and heparan sulfate/ heparin even within a single polymer chain (Esko and Lindahl, 2001). A few clever microbes also produce GAG chains, but sulfation or epimerization have not been yet described. The chondroitin and heparan sulfate/heparin chains in vertebrates are initially synthesized by elongation of a xylose-containing linkage tetrasaccharide attached to a variety of proteins. Keratan is either O-linked or N-linked to certain proteins depending on the particular molecule. HA and all of the known bacterial GAGs are not part of glycoproteins. This review focuses on the identity and the nature of the known microbial glycosyltransferases that produce GAG chains (Table II). Basics of GAG glycosyltransferases All of the known HA, chondroitin, and heparan sulfate/heparin glycosyltransferase enzymes that synthesize the alternating sugar repeat backbones in microbes and in vertebrates utilize uridine diphospho-(UDP)-sugar precursors and metal cofactors (e.g., magnesium and/or manganese ion) near neutral pH according to the overall reaction: n UDP-GlcUA + n UDP-HexNAc → 2n UDP + [GlcUA-HexNAc]n where HexNAc = GlcNAc or GalNAc. Depending on the specific GAG and the particular organism or tissue examined, the degree of polymerization, n, ranges from ∼25 to ∼10,000. The bacterial GAG glycosyltransferase polypeptides are associated with the cell membranes; this localization makes sense with respect to synthesis of polysaccharide molecules destined for the cell surface. Various names for the GAG glycosyltransferases have been used in the literature over the last four decades. The dual-action enzymes required for the production of the HA chain have been called synthases (or, in early reports, synthetases). The enzymes that elongate the repeating chondroitin or the repeating heparan sulfate/heparin backbone have been called various names, including co-polymerases, co-transferases, polymerases, synthases, or the individual component activities 9R P.L. De Angelis Table I. Structures of the vertebrate and microbial GAG repeating backbones Postpolymerization modifications Polymer Disaccharide repeat Vertebrates Bacteria Hyaluronan β3GlcNAcβ4GlcUA none none Chondroitin β3GalNAcβ4GlcUA O-sulfated, epimerized none or fructose( β1,3)GlcUA Heparan sulfate/ heparin α4GlcNAcβ4GlcUA O-, N-sulfated, epimerized none Keratan β4GlcNAcβ3Gal O-sulfated not reported Table II. Microbes, diseases, GAGs, and glycosyltransferases Bacteria (disease) a Hyaluronan Chondroitin Heparosan Enzyme [sizeb]/GenBank Streptococcus Group A (pharyngitis, scarlet fever, necrotizing fascitis, impetigo, and more) X spHAS [419]/L20853 Group C (mastitis*, strangles*) X seHAS [418]/AF023876 Escherichia coli Xc K4 (diarrhea) K5 (urinary tract and extraintestinal infections, sepsis) not reported X Kfi A [238] + KfiC [520] complex/X77617 Pasteurella multocida Type A (fowl cholera*, shipping fever*, sepsis from animal bites) X pmHAS [972]/AF036004 Type D (atrophic rhinitis*) Type F (fowl cholera*) X X pmHS [617]/AF425591 pmCS [965]/AF195517 aAnimal diseases denoted with a asterisk (remainder are human diseases). bNumber of amino acid residues in the deduced open reading frame. cFructosylated polymer. were directly termed (e.g., glucuronic acid [GlcUA]-transferase, GlcNAc-transferase, or GalNAc-transferase). Hyaluronan synthases of Streptococcus and Pasteurella The HA extracellular capsules of Gram-positive Group A Streptococcus (Kendall et al., 1937) and Gram-negative Type A Pasteurella multocida (Carter and Annau, 1953) were shown to be identical to HA of vertebrates. As the vertebrate HA synthases (HASs) were (and remain) relatively difficult to study biochemically, more initial progress was made on the “simpler,” higher specific activity membrane preparations of str (...truncated)