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Polysaccharide Lyase Family 15

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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

Substrate specificities

PL15 contains 2 subfamilies [1] as well as several proteins not assigned to any subfamily. Subfamily 1 has been shown to only degrade alginate [2, 3, 4, 5] while subfamily 2 has been found to be heparin and heparan sulfate lyases [6, 7]. Alginate consist 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 [8, 9]. Heparin consist 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 [10].

Kinetics and Mechanism

Figure 1 +1 subsite of the alginate lyase Atu3025 with the MGG substrate.

The β-elimination catalyzed by the PL15 enzymes results in the formation of a C4-C5 unsaturated sugar residue at the new non-reducing end. The first step is the neutralization of the acid group in the +1 subsite by the conserved H531 and R314 (Atu3025 numbering)[3]. This lowers the pKa value of the C5-proton allowing for abstraction by the catalytic base (Figure 1). A catalytic acid then donates a proton to the glycosidic linkage resulting in the β-elimination.

Catalytic Residues

After charge neutralization a histidine functions as the catalytic base and a tyrosine as the acid. They were originally identified as H311 and Y365 in Atu3025 from Agrobacterium fabrum [3].

Three-dimensional structures

Figure 2 Crystal structures of the substrate complex of the monomeric Atu3025 (PDB ID 3AFL) with the MGG substrate in blue.

The first crystal structure available for a PL15 member was that of the alginate lyase Atu3025 from Agrobacterium fabrum (Figure 2) [3]. The catalytic domains consists of an N-terminal (α/α)6 barrel domain and a C-terminal anti-parallel β-sheet domain. The catalytic site is located between the two domains with the catalytic residues and the arginine charge neutralizer located in the (α/α)6 barrel and the histidine neutralizer in a loop extending into the active site from the anti-parallel β-sheet domain [3].

Family Firsts

First catalytic activity
Alginate lyase IV from Sphingomonas sp activity shown against alginate di- and trisaccharides by TLC from purified protein [2].
First catalytic base/acid
Atu3025 from Agrobacterium fabrum. H311 and Y365 was suggested as acid/base based upon the crystal structure of the substrate complex, residue conservation, mutagenesis and activity analysis (H311A: inactive and Y365A: 0.3 % activity remaining)[3].
First charge neutralizer
Atu3025 from Agrobacterium fabrum H531 was suggested based on the crystal structure, its conservation, mutagenesis and activity analysis (H531A 0.45 % activity). R314 is proposed based on its proximity to the carboxylate group in the +1 subsite and its conservation [3].
First 3-D structure
Atu3025 from Agrobacterium fabrum an exo alginate lyase from subfamily 1 [3].


  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 | PubMed ID:20925655 [Lombard2010]
  2. 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 | PubMed ID:12729723 [Miyake2003]
  3. 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 | PubMed ID:20507980 [Ochiai2010]
  4. 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 | PubMed ID:24795372 [Jagtap2014]
  5. 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 | PubMed ID:16233753 [Hashimoto20005]
  6. 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 | PubMed ID:28630303 [Cartmell2017]
  7. 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 | PubMed ID:30850540 [Helbert2019]
  8. Haug, A., Larsen, B., and Smidsrod, O. (1967) Studies on sequence of uronic acid residues in alginic acid. Acta Chem. Scand. 21, 691–704. DOI:10.3891/acta.chem.scand.21-0691

  9. Haug, A., Larsen, B., and Smidsrod, O. (1966) A study of constitution of alginic acid by partial acid hydrolysis. Acta Chem. Scand. 20, 183–190. DOI:10.3891/acta.chem.scand.20-0183

  10. Garron ML and Cygler M. (2010). Structural and mechanistic classification of uronic acid-containing polysaccharide lyases. Glycobiology. 2010;20(12):1547-73. DOI:10.1093/glycob/cwq122 | PubMed ID:20805221 [Garron2010]

All Medline abstracts: PubMed