Polysaccharide epimerases

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• Author: ^^^Margrethe Gaardlos^^^ and ^^^Anne Tondervik^^^
• Responsible Curator: ^^^Finn Aachmann^^^

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Mannuronan C5-epimerases

Substrate specificity

Mannuronan C5-epimerases are a group of enzymes that catalyze epimerization at the polymer-level of β-d-mannuronic acid residues (hereafter denoted M) into α-l-guluronic acid residues (hereafter denoted G) in alginate [1, 2, 3]. Alginate is an anionic polysaccharide made by brown seaweeds, some species of red algae, and the gram-negative bacterial genera Pseudomonas and Azotobacter [4, 5, 6, 7, 8]. The function of alginate in the different organisms are various, and related to structure, protection and surface adhesion [9, 10, 11, 12]. Alginate is a copolymer of the two 1-4 linked epimers [13, 14, 15], and by changing the composition of the two monomers the epimerases fine-tune the properties of the polymer [16].

At first, alginate is made as a homopolymer of M in the cell. Epimerases then convert some of the M residues in the polymer into G-residues [3, 17, 18]. This epimerization is not random and creates block structures of M, G or alternating MG [19, 20]. Alginate residues that are oxidized or acetylated are not substrates for the epimerases, and acetylation of alginate could be a way to control epimerization in nature [21, 22].

A. Haworth structures of d-mannuronic acid (M) and l-guluronic acid (G). B. The three different block structures found in alginate: non-epimerized M-blocks, MG-blocks and G-blocks.

Mannuronan C5-epimerases exist both in algae and in bacteria [1, 23]. Gene analyses propose as many as 31 different genes encoding putative mannuronan C-5 epimerases in the brown algae Ectocarpus [24]. However, the algal epimerases are difficult to express and it is the bacterial enzymes that have been studied most extensively [24, 25]. Two categories of bacterial mannuronan C-5-epimerases have been described: the periplasmic AlgG and the extracellular and calcium dependent AlgE. AlgG creates single G residues in stretches of mannuronan, while the AlgE enzymes are processive and create MG-blocks and G-blocks. Pseudomonas is only known to produce AlgG [18, 26, 27], while A. vinelandii contains seven active AlgE enzymes in addition to AlgG [28, 29, 30, 31]. A mutant strain of P. fluorescens without the algG gene creates pure mannuronan [32]. This strain can be used to produce unepimerized substrate, which is useful for the study of the epimerization reaction. Methods for studying this are discussed in a later section.

References

1. Haug A and Larsen B. (1969). Biosynthesis of alginate. Epimerisation of D-mannuronic to L-guluronic acid residues in the polymer chain. Biochim Biophys Acta. 1969;192(3):557-9. DOI:10.1016/0304-4165(69)90414-0 |
2. Larsen B and Haug A. (1971). Biosynthesis of alginate. 1. Composition and structure of alginate produced by Azotobacter vinelandii (Lipman). Carbohydr Res. 1971;17(2):287-96. DOI:10.1016/s0008-6215(00)82536-7 |
3. Haug A and Larsen B. (1971). Biosynthesis of alginate. II. Polymannuronic acid C-5-epimerase from Azotobacter vinelandii (Lipman). Carbohydr Res. 1971;17(2):297-308. DOI:10.1016/s0008-6215(00)82537-9 |

4. Stanford, Edw C C. (1883) On algin: a new substance obtained from some of the commoner species of marine algae. R. Anderson. NLM ID: 101217546

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8. Okazaki, M., K. and Furuya, K. Tsukayam and K. Nisizawa. (1982) Isolation and Identification of Alginic Acid from a Calcareous Red Alga Serraticardia maxima. Botanica Marina, vol. 25, no. 3., pp. 123-131. [1]

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19. Haug, Arne and Larsen, Bjørn and Smidsrød, Olav. (1966) A study on the constitution of alginic acidby partial acid hydrolysis. Acta Chemica Scandinavica, vol. 5 (July), pp. 271–277. [1]

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22. Kristiansen, Kåre A and Schirmer, Bjørn C and Aachmann, Finn L. and Skjåk-Bræk, Gudmund and Draget, Kurt I. and Christensen, Bjørn E. (2009) Novel alginates prepared by independent control of chain stiff-ness and distribution of G-residues: Structure and gelling properties. Carbohydrate Polymers, vol. 77, no.4, pp. 725–735. [1]

23. Madgwick J, Haug A, and Larsen B. (1973). Polymannuronic acid 5-epimerase from the marine alga Pelvetia canaliculata (L.) Dcne. et Thur. Acta Chem Scand. 1973;27(9):3592-4. DOI:10.3891/acta.chem.scand.27-3592 |
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27. Morea A, Mathee K, Franklin MJ, Giacomini A, O'Regan M, and Ohman DE. (2001). Characterization of algG encoding C5-epimerase in the alginate biosynthetic gene cluster of Pseudomonas fluorescens. Gene. 2001;278(1-2):107-14. DOI:10.1016/s0378-1119(01)00685-0 |
28. Ertesvåg H, Doseth B, Larsen B, Skjåk-Braek G, and Valla S. (1994). Cloning and expression of an Azotobacter vinelandii mannuronan C-5-epimerase gene. J Bacteriol. 1994;176(10):2846-53. DOI:10.1128/jb.176.10.2846-2853.1994 |
29. Ertesvåg H, Høidal HK, Hals IK, Rian A, Doseth B, and Valla S. (1995). A family of modular type mannuronan C-5-epimerase genes controls alginate structure in Azotobacter vinelandii. Mol Microbiol. 1995;16(4):719-31. DOI:10.1111/j.1365-2958.1995.tb02433.x |
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