Polysaccharide Lyase Family 7

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• Authors: ^^^Nadine Gerlach^^^ and ^^^Jan-Hendrik Hehemann^^^
• Responsible Curator: ^^^Jan-Hendrik Hehemann^^^

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Polysaccharide Lyase Family PL7
3D Structure	β jelly roll
Mechanism	β-elimination
Active site residues	known
CAZy DB link
https://www.cazy.org/PL7.html

Mechanism

Alginate lyases (Alys) of all families, PL5-7, PL14, PL15, PL17-18, cleave the glycosidic bond via β-elimination. Most PL7s are endo-active, i.e. acting within a poly- or oligosaccharide and releasing smaller alginate fragments. exo activity [1]

In contrast to terrestrial PL7, marine PLs need the divalent cation calcium for substrate recognition and binding [1]. Ca2+ is weakening the ionic interactions between substrate (polyanion) and PL7 (polycation) by reducing the surface density of the alginate charge and therefore increasing the enzyme activity [2].

Kinetics and catalytic residues

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Figure 1. Multiple protein alignment of Aly PL7 as well as a secondary structure prediction of the crystallized PL7 from Klabsiella pneumoniae (PDB code: 4OXZ, [3]). Conserved residues in the homologues are colored in red and (putative) catalytic residues are indicated by a star.

Arg (R), Gln (Q), His (H), Tyr (Y) form active site [4] His removes negative charge of the substrate additional negatively charged residues [1]

Substrate specificities

The polysaccharide lyase family 7 (PL7) contains five subfamilies (SF) based on their sequence similiarities [5], plus a so far uncharacterized sixth subfamily, which sonstist only of marine representatives of the Flavobacteriaceae [1]. All characterized PL7 enzymes were alginate lyases specific for the anionic, gel forming polysaccharide alginate. The substrate specificity depends on the source of alginate, i.e. derived from brown seaweed or mucoid bacteria Pseudomonas spp. and Azotobacter vinelandii, as well as geographical and saisonal parameters. Alginate is an heteropolysaccharide, consisting of β-D-mannuronate (M) and α-L-guluronate (G). These monosaccharides can occur in homogenous and heterogenous blocks. Hence, PL7 lyases can be mannuronate (EC 4.2.2.3), guluronate (EC 4.2.2.11) or mixed link (EC 4.2.2.-) specific. Despite the pefernce for M- or G-enriched blocks, most PL7s also have a moderate to low processivity for the other building block [1], [6].

SF3 & SF5 G-specific [1]

Substitution of hydrophobic amino acids in the isoleucine site of domain QIH could have an enormous influence on the high-affinity to pM or pG. This isoleucine was reconfirmed to be indispensable for recognition of the pG or G-G bond [7]

Three-dimensional structures

Figure 2. 3D Structure of endo- and exo-active PL7s [1]. (A,C) AlyA1 and (B, D) AlyA5 from Zobellia galaactinovorans DsijT shown as cartoon (A,C) and surface structure (B,D) with superimposed tetrasaccharide from PDB:2ZAA [8]

PL 7 is a very well biochemical characterized family with almost 40 entries in the CAZy data base [9]. Structural insights on the other hand are still restricted with nine 3D structures from only eight bacterial strains (status at CAZy in August 2019). The first structure of a PL7 was determined from Pseudomonas aeruginosa by multiple isomorphous replacement (MIR) at 2.0 Å resolution [10]. Just like PL14, PL7 belongs to the jelly roll family with a wide open cleft harboring the active site. Til date, there is only one known exoactive PL7 structure. Zobellia galaactinovorans DsijT is harboring, among others, two PL7 with two completely different activity modifs. AlyA1 belongs to SF3 and is an endo-active PL7, which active site . AlyA5 belongs to SF5 and is exo-active, which active site is close by three additional loops forming a small pocket.

[1]

CBM32

Evolution of Aly PULs

lyases play different roles and have complementary activities [1] [11]

Family Firsts

First catalytic endo-activity

First catalytic exo-activity

AlyA5 from Zobellia galactanivorans DsijT [1]

First 3-D apo-structure

PA1167 from Pseudomonas aeruginosa [10]

First 3-D holo-structure

A1-II from Sphingomons sp. A1 [8]

References

1. Thomas F, Lundqvist LC, Jam M, Jeudy A, Barbeyron T, Sandström C, Michel G, and Czjzek M. (2013). Comparative characterization of two marine alginate lyases from Zobellia galactanivorans reveals distinct modes of action and exquisite adaptation to their natural substrate. J Biol Chem. 2013;288(32):23021-37. DOI:10.1074/jbc.M113.467217 |
2. Favorov VV, Vozhova EI, Denisenko VA, and Elyakova LA. (1979). A study of the reaction catalysed by alginate lyase VI from the sea mollusc, Littorina sp. Biochim Biophys Acta. 1979;569(2):259-66. DOI:10.1016/0005-2744(79)90061-5 |

3. Crystal Structure of the alginate lyase from Klebsiella pneumoniae

4. Yamasaki M, Ogura K, Hashimoto W, Mikami B, and Murata K. (2005). A structural basis for depolymerization of alginate by polysaccharide lyase family-7. J Mol Biol. 2005;352(1):11-21. DOI:10.1016/j.jmb.2005.06.075 |
5. 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 |
6. Badur AH, Jagtap SS, Yalamanchili G, Lee JK, Zhao H, and Rao CV. (2015). Alginate lyases from alginate-degrading Vibrio splendidus 12B01 are endolytic. Appl Environ Microbiol. 2015;81(5):1865-73. DOI:10.1128/AEM.03460-14 |
7. Deng S, Ye J, Xu Q, and Zhang H. (2014). Structural and functional studies on three alginate lyases from Vibrio alginolyticus. Protein Pept Lett. 2014;21(2):179-87. DOI:10.2174/09298665113206660094 |

8. Crystal Structure of Family 7 Alginate Lyase A1-II' H191N/Y284F in Complex with Substrate (GGMG)

9. Lombard V, Golaconda Ramulu H, Drula E, Coutinho PM, and Henrissat B. (2014). The carbohydrate-active enzymes database (CAZy) in 2013. Nucleic Acids Res. 2014;42(Database issue):D490-5. DOI:10.1093/nar/gkt1178 |
10. Yamasaki M, Moriwaki S, Miyake O, Hashimoto W, Murata K, and Mikami B. (2004). Structure and function of a hypothetical Pseudomonas aeruginosa protein PA1167 classified into family PL-7: a novel alginate lyase with a beta-sandwich fold. J Biol Chem. 2004;279(30):31863-72. DOI:10.1074/jbc.M402466200 |
11. Hehemann JH, Correc G, Barbeyron T, Helbert W, Czjzek M, and Michel G. (2010). Transfer of carbohydrate-active enzymes from marine bacteria to Japanese gut microbiota. Nature. 2010;464(7290):908-12. DOI:10.1038/nature08937 |