Polysaccharide Lyase Family 15

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• Author: ^^^Emil Stender^^^
• Responsible Curator: ^^^Birte Svensson^^^

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Polysaccharide Lyase Family 15
3D structure	(α/α)6 barrel + anti-parallel β-sheet
Mechanism	β-elimination
Charge neutralizer	Arginine and histidine
Active site residues	known
CAZy DB link
https://www.cazy.org/PL15.html

Substrate specificities

PL15 currently contains 2 subfamilies [1] as well as several proteins currently not assigned to any subfamily. Subfamily 1 has been shown to only degrade alginate ADDIN CSL_CITATION {"citationItems":[{"id":"ITEM-1","itemData":{"DOI":"10.1016/S1046-5928(03)00018-4","ISSN":"10465928","PMID":"12729723","abstract":"Sphingomonas sp. A1 (strain A1) cells contain three kinds of endotype alginate lyases [A1-I, A1-II, and A1-III], all of which are formed from a common precursor through posttranslational processing. In addition to these lyases, another type of lyase (A1-IV) that acts on oligoalginates exists in the bacterium. A1-IV was overexpressed in Escherichia coli cells through control of its gene under the T7 promoter. The expression level of the enzyme in E. coli cells was 8.6 U/L-culture, which was about 270-fold higher than that in strain A1 cells. The enzyme was purified to homogeneity through three steps with an activity yield of 10.9%. The optimal pH and temperature, thermal stability, and mode of action of the purified enzyme were similar to those of the native enzyme from strain A1 cells. A1-IV exolytically degraded oligoalginates, which were produced from alginate through the reaction of A1-I, A1-II, or A1-III, into monosaccharides, indicating that the cooperative actions of these four enzymes cause the complete depolymerization of alginate in strain A1 cells. © 2003 Elsevier Science (USA). All rights reserved.","author":[{"dropping-particle":"","family":"Miyake","given":"Osamu","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Hashimoto","given":"Wataru","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Murata","given":"Kousaku","non-dropping-particle":"","parse-names":false,"suffix":""}],"container-title":"Protein Expression and Purification","id":"ITEM-1","issue":"1","issued":{"date-parts":"2003"},"page":"33-41","title":"An exotype alginate lyase in Sphingomonas sp. A1: overexpression in Escherichia coli, purification, and characterization of alginate lyase IV (A1-IV)","type":"article-journal","volume":"29"},"uris":["http://www.mendeley.com/documents/?uuid=a10b46a4-bdd2-4166-90b1-4a4c7c369462%22]},{"id":"ITEM-2","itemData":{"DOI":"10.1074/jbc.M110.125450","ISBN":"0021-9258","ISSN":"1083351X","PMID":"20507980","abstract":"Alginate, a major component of the cell wall matrix in brown seaweeds, is degraded by alginate lyases through a beta-elimination reaction. Almost all alginate lyases act endolytically on substrate, thereby yielding unsaturated oligouronic acids having 4-deoxy-l-erythro-hex-4-enepyranosyluronic acid at the nonreducing end. In contrast, Agrobacterium tumefaciens alginate lyase Atu3025, a member of polysaccharide lyase family 15, acts on alginate polysaccharides and oligosaccharides exolytically and releases unsaturated monosaccharides from the substrate terminal. The crystal structures of Atu3025 and its inactive mutant in complex with alginate trisaccharide (H531A/DeltaGGG) were determined at 2.10- and 2.99-A resolutions with final R-factors of 18.3 and 19.9%, respectively, by x-ray crystallography. The enzyme is comprised of an alpha/alpha-barrel + anti-parallel beta-sheet as a basic scaffold, and its structural fold has not been seen in alginate lyases analyzed thus far. The structural analysis of H531A/DeltaGGG and subsequent site-directed mutagenesis studies proposed the enzyme reaction mechanism, with His(311) and Tyr(365) as the catalytic base and acid, respectively. Two structural determinants, i.e. a short alpha-helix in the central alpha/alpha-barrel domain and a conformational change at the interface between the central and C-terminal domains, are essential for the exolytic mode of action. This is, to our knowledge, the first report on the structure of the family 15 enzyme.","author":[{"dropping-particle":"","family":"Ochiai","given":"Akihito","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Yamasaki","given":"Masayuki","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Mikami","given":"Bunzo","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Hashimoto","given":"Wataru","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Murata","given":"Kousaku","non-dropping-particle":"","parse-names":false,"suffix":""}],"container-title":"Journal of Biological Chemistry","id":"ITEM-2","issue":"32","issued":{"date-parts":"2010"},"page":"24519-24528","title":"Crystal Structure of exotype alginate lyase Atu3025 from Agrobacterium tumefaciens","type":"article-journal","volume":"285"},"uris":["http://www.mendeley.com/documents/?uuid=68ef39c9-91a5-484b-b750-19b4460ae650%22]},{"id":"ITEM-3","itemData":{"DOI":"10.1128/AEM.01285-14","ISSN":"10985336","PMID":"24795372","abstract":"Marine microbes use alginate lyases to degrade and catabolize alginate, a major cell wall matrix polysaccharide of brown seaweeds. Microbes frequently contain multiple, apparently redundant alginate lyases, raising the question of whether these enzymes have complementary functions. We report here on the molecular cloning and functional characterization of three exo-type oligoalginate lyases (OalA, OalB, and OalC) from Vibrio splendidus 12B01 (12B01), a marine bacterioplankton species. OalA was most active at 16°C, had a pH optimum of 6.5, and displayed activities toward poly-β-d-mannuronate [poly(M)] and poly-α-l-guluronate [poly(G)], indicating that it is a bifunctional enzyme. OalB and OalC were most active at 30 and 35°C, had pH optima of 7.0 and 7.5, and degraded poly(M·G) and poly(M), respectively. Detailed kinetic analyses of oligoalginate lyases with poly(G), poly(M), and poly(M·G) and sodium alginate as substrates demonstrated that OalA and OalC preferred poly(M), whereas OalB preferred poly(M·G). The catalytic efficiency (kcat/Km) of OalA against poly(M) increased with decreasing size of the substrate. OalA showed kcat/Km from 2,130 mg(-1) ml s(-1) for the trisaccharide to 224 mg(-1) ml s(-1) for larger oligomers of ∼50 residues, and 50.5 mg(-1) ml s(-1) for high-molecular-weight alginate. Although OalA was most active on the trisaccharide, OalB and OalC preferred dimers. Taken together, our results indicate that these three Oals have complementary substrate scopes and temperature and pH adaptations.","author":[{"dropping-particle":"","family":"Jagtap","given":"Sujit Sadashiv","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Hehemann","given":"Jan Hendrik","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Polz","given":"Martin F.","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Lee","given":"Jung Kul","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Zhao","given":"Huimin","non-dropping-particle":"","parse-names":false,"suffix":""}],"container-title":"Applied and Environmental Microbiology","id":"ITEM-3","issue":"14","issued":{"date-parts":"2014"},"page":"4207-4214","title":"Comparative biochemical characterization of three exolytic oligoalginate lyases from Vibrio splendidus reveals complementary substrate scope, temperature, and pH adaptations","type":"article-journal","volume":"80"},"uris":["http://www.mendeley.com/documents/?uuid=dcde9ba3-b1cf-4f7d-b9a8-ebdcd84a07fa%22]},{"id":"ITEM-4","itemData":{"DOI":"10.1263/jbb.99.048","ISSN":"1389-1723","PMID":"16233753","abstract":"Sphingomonas sp. A1 (strain A1) produces three endotypes (A1-I [65 kDa], A1-II [25 kDa], and A1-III [40 kDa]) and an exotype (A1-IV [86 kDa]) alginate lyases in cytoplasm. These four enzymes cooperatively depolymerize alginate into constituent monosaccharides. In addition to the genes for these lyases, novel genes encoding hypothetical proteins homologous with A1-IV were found in the genomes of many bacteria including strain A1. One such protein, A1-IV' (90 kDa) of strain A1, was overexpressed in Escherichia coli cells, purified, and characterized. A1-IV' catalyzed the cleavage of glycosidic bonds in alginate through a beta-elimination reaction and released unsaturated di- and trisaccharides as main products, thus indicating that the enzyme is an endotype alginate lyase. A1-IV', which differed from A1-IV in some enzymatic properties, was not expressed in strain A1, suggesting that A1-IV' has no significant role in alginate metabolism. A1-IV' and other A1-IV homologs facilitate the creation of novel polysaccharide lyase family 15 based on their primary structures, implying the evolution route of alginate lyases in family PL-15.","author":[{"dropping-particle":"","family":"Hashimoto","given":"Wataru","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Miyake","given":"Osamu","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Ochiai","given":"Akihito","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Murata","given":"Kousaku","non-dropping-particle":"","parse-names":false,"suffix":""}],"container-title":"Journal of bioscience and bioengineering","id":"ITEM-4","issue":"1","issued":{"date-parts":"2005"},"page":"48-54","title":"Molecular identification of Sphingomonas sp. A1 alginate lyase (A1-IV') as a member of novel polysaccharide lyase family 15 and implications in alginate lyase evolution.","type":"article-journal","volume":"99"},"uris":["http://www.mendeley.com/documents/?uuid=51cad4de-8e15-475c-acec-a6942bed1400%22]}],"mendeley":{"formattedCitation":"(2–5)","plainTextFormattedCitation":"(2–5)","previouslyFormattedCitation":"(2–5)"},"properties":{"noteIndex":0},"schema":"https://github.com/citation-style-language/schema/raw/master/csl-citation.json%22}(2–5) while subfamily 2 has been found to be heparin and heparan sulfate lyases ADDIN CSL_CITATION {"citationItems":[{"id":"ITEM-1","itemData":{"DOI":"10.1073/pnas.1704367114","ISSN":"0027-8424","PMID":"28630303","abstract":"The human microbiota, which plays an important role in health and disease, uses complex carbohydrates as a major source of nutrients. Utilization hierarchy indicates that the host glycosaminoglycans heparin (Hep) and heparan sulfate (HS) are high-priority carbohydrates for Bacteroides thetaiotaomicron, a prominent member of the human microbiota. The sulfation patterns of these glycosaminoglycans are highly variable, which presents a significant enzymatic challenge to the polysaccharide lyases and sulfatases that mediate degradation. It is possible that the bacterium recruits lyases with highly plastic specificities and expresses a repertoire of enzymes that target substructures of the glycosaminoglycans with variable sulfation or that the glycans are desulfated before cleavage by the lyases. To distinguish between these mechanisms, the components of the B. thetaiotaomicron Hep/HS degrading apparatus were analyzed. The data showed that the bacterium expressed a single-surface endo-acting lyase that cleaved HS, reflecting its higher molecular weight compared with Hep. Both Hep and HS oligosaccharides imported into the periplasm were degraded by a repertoire of lyases, with each enzyme displaying specificity for substructures within these glycosaminoglycans that display a different degree of sulfation. Furthermore, the crystal structures of a key surface glycan binding protein, which is able to bind both Hep and HS, and periplasmic sulfatases reveal the major specificity determinants for these proteins. The locus described here is highly conserved within the human gut Bacteroides, indicating that the model developed is of generic relevance to this important microbial community.","author":[{"dropping-particle":"","family":"Cartmell","given":"Alan","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Lowe","given":"Elisabeth C.","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Baslé","given":"Arnaud","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Firbank","given":"Susan J.","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Ndeh","given":"Didier A.","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Murray","given":"Heath","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Terrapon","given":"Nicolas","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Lombard","given":"Vincent","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Henrissat","given":"Bernard","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Turnbull","given":"Jeremy E.","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Czjzek","given":"Mirjam","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Gilbert","given":"Harry J.","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Bolam","given":"David N.","non-dropping-particle":"","parse-names":false,"suffix":""}],"container-title":"Proceedings of the National Academy of Sciences","id":"ITEM-1","issue":"27","issued":{"date-parts":"2017"},"page":"7037-7042","title":"How members of the human gut microbiota overcome the sulfation problem posed by glycosaminoglycans","type":"article-journal","volume":"114"},"uris":["http://www.mendeley.com/documents/?uuid=338ab035-66d5-4197-9ce8-779b9805b183%22]},{"id":"ITEM-2","itemData":{"DOI":"10.1073/pnas.1815791116","ISSN":"0027-8424","abstract":"Over the last two decades, the number of gene/protein sequences gleaned from sequencing projects of individual genomes and environmental DNA has grown exponentially. Only a tiny fraction of these predicted proteins has been experimentally characterized, and the function of most proteins remains hypothetical or only predicted based on sequence similarity....","author":[{"dropping-particle":"","family":"Helbert","given":"William","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Poulet","given":"Laurent","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Drouillard","given":"Sophie","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Mathieu","given":"Sophie","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Loiodice","given":"Mélanie","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Couturier","given":"Marie","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Lombard","given":"Vincent","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Terrapon","given":"Nicolas","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Turchetto","given":"Jeremy","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Vincentelli","given":"Renaud","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Henrissat","given":"Bernard","non-dropping-particle":"","parse-names":false,"suffix":""}],"container-title":"Proceedings of the National Academy of Sciences of the United States of America","id":"ITEM-2","issue":"13","issued":{"date-parts":"2019"},"page":"6063-6068","title":"Discovery of novel carbohydrate-active enzymes through the rational exploration of the protein sequences space","type":"article-journal","volume":"116"},"uris":["http://www.mendeley.com/documents/?uuid=e0b9a3a0-aadb-49af-ad30-b5ef4878a6df%22]}],"mendeley":{"formattedCitation":"(6, 7)","plainTextFormattedCitation":"(6, 7)","previouslyFormattedCitation":"(6, 7)"},"properties":{"noteIndex":0},"schema":"https://github.com/citation-style-language/schema/raw/master/csl-citation.json%22}(6, 7). Alginate consisting of 1,4 linked β-D-mannuronic acid and α-L-guluronic acid arranged in poly-mannuronic acid blocks, poly-guluronic acid blocks or poly-mannuronic/guluronic acid blocks ADDIN CSL_CITATION {"citationItems":[{"id":"ITEM-1","itemData":{"DOI":"10.3891/acta.chem.scand.21-0691","ISSN":"0904-213X","author":[{"dropping-particle":"","family":"Haug","given":"A","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Larsen","given":"B","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Smidsrod","given":"O","non-dropping-particle":"","parse-names":false,"suffix":""}],"container-title":"Acta Chemica Scandinavica","id":"ITEM-1","issue":"3","issued":{"date-parts":"1967"},"page":"691-704","title":"Studies on sequence of uronic acid residues in alginic acid","type":"article-journal","volume":"21"},"uris":["http://www.mendeley.com/documents/?uuid=bbbac0a0-1a50-4d81-91fb-b5ea25ba38b7%22]},{"id":"ITEM-2","itemData":{"DOI":"10.3891/acta.chem.scand.20-0183","ISSN":"0904-213X","author":[{"dropping-particle":"","family":"Haug","given":"A","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Larsen","given":"B","non-dropping-particle":"","parse-names":false,"suffix":""},{"dropping-particle":"","family":"Smidsrod","given":"O","non-dropping-particle":"","parse-names":false,"suffix":""}],"container-title":"Acta Chemica Scandinavica","id":"ITEM-2","issue":"1","issued":{"date-parts":"1966"},"page":"183-190","title":"A study of constitution of alginic acid by partial acid hydrolysis","type":"article-journal","volume":"20"},"uris":["http://www.mendeley.com/documents/?uuid=e8a81ee7-106f-4eef-a522-8f1ce49ea0a7%22]}],"mendeley":{"formattedCitation":"(8, 9)","plainTextFormattedCitation":"(8, 9)","previouslyFormattedCitation":"(8, 9)"},"properties":{"noteIndex":0},"schema":"https://github.com/citation-style-language/schema/raw/master/csl-citation.json%22}(8, 9). Heparin consisting of disaccharide repeating units of which the most common is 2-O-sulfated 1,4 linked α-L-iduronic acid and 6-O-sulfated, N-sulfated glucosamine [IdoA(2S)-GlcNS(6S)]. Heparan sulfate being very similar to heparin having the IdoA replaced with β-D-glucuronic acid with a considerably more variable sulfation and acetylation pattern. Authors may get an idea of what to put in each field from Curator Approved Glycoside Hydrolase Families. (TIP: Right click with your mouse and open this link in a new browser window...)

In the meantime, please see these references for an essential introduction to the CAZy classification system: [2, 3].

Kinetics and Mechanism

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Catalytic Residues

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Three-dimensional structures

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Family Firsts

First stereochemistry determination

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First catalytic nucleophile identification

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First general acid/base residue identification

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First 3-D structure

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References

1. Lombard V, Bernard T, Rancurel C, Brumer H, Coutinho PM, and Henrissat B. (2010). A hierarchical classification of polysaccharide lyases for glycogenomics. Biochem J. 2010;432(3):437-44. DOI:10.1042/BJ20101185 |

2. Davies, G.J. and Sinnott, M.L. (2008) Sorting the diverse: the sequence-based classifications of carbohydrate-active enzymes. The Biochemist, vol. 30, no. 4., pp. 26-32. Download PDF version.

3. Cantarel BL, Coutinho PM, Rancurel C, Bernard T, Lombard V, and Henrissat B. (2009). The Carbohydrate-Active EnZymes database (CAZy): an expert resource for Glycogenomics. Nucleic Acids Res. 2009;37(Database issue):D233-8. DOI:10.1093/nar/gkn663 |
4. Miyake O, Hashimoto W, and Murata K. (2003). An exotype alginate lyase in Sphingomonas sp. A1: overexpression in Escherichia coli, purification, and characterization of alginate lyase IV (A1-IV). Protein Expr Purif. 2003;29(1):33-41. DOI:10.1016/s1046-5928(03)00018-4 |
5. Ochiai A, Yamasaki M, Mikami B, Hashimoto W, and Murata K. (2010). Crystal structure of exotype alginate lyase Atu3025 from Agrobacterium tumefaciens. J Biol Chem. 2010;285(32):24519-28. DOI:10.1074/jbc.M110.125450 |
6. Jagtap SS, Hehemann JH, Polz MF, Lee JK, and Zhao H. (2014). Comparative biochemical characterization of three exolytic oligoalginate lyases from Vibrio splendidus reveals complementary substrate scope, temperature, and pH adaptations. Appl Environ Microbiol. 2014;80(14):4207-14. DOI:10.1128/AEM.01285-14 |
7. Hashimoto W, Miyake O, Ochiai A, and Murata K. (2005). Molecular identification of Sphingomonas sp. A1 alginate lyase (A1-IV') as a member of novel polysaccharide lyase family 15 and implications in alginate lyase evolution. J Biosci Bioeng. 2005;99(1):48-54. DOI:10.1263/jbb.99.48 |
8. Cartmell A, Lowe EC, Baslé A, Firbank SJ, Ndeh DA, Murray H, Terrapon N, Lombard V, Henrissat B, Turnbull JE, Czjzek M, Gilbert HJ, and Bolam DN. (2017). How members of the human gut microbiota overcome the sulfation problem posed by glycosaminoglycans. Proc Natl Acad Sci U S A. 2017;114(27):7037-7042. DOI:10.1073/pnas.1704367114 |
9. Helbert W, Poulet L, Drouillard S, Mathieu S, Loiodice M, Couturier M, Lombard V, Terrapon N, Turchetto J, Vincentelli R, and Henrissat B. (2019). Discovery of novel carbohydrate-active enzymes through the rational exploration of the protein sequences space. Proc Natl Acad Sci U S A. 2019;116(13):6063-6068. DOI:10.1073/pnas.1815791116 |