Structural insights into marine carbohydrate degradation by family GH16 kappa-carrageenases.

4 years ago

Structural insights into marine carbohydrate degradation by family GH16 kappa-carrageenases.

Tristan Barbeyron, Cédric Leroux, Murielle Jam, Robert Larocque, Thomas Bernard, Gurvan Michel, Aurélie Préchoux, Alexandra Jeudy, Maria Matard-Mann, Mirjam Czjzek, Pi Nyvall-Collén

Carrageenans are sulfated α-1,3-β-1,4-galactans found in the cell wall of some red algae that are practically valuable for their gelation and biomimetic properties but also serve as a potential carbon source for marine bacteria. Carbohydrate degradation has been studied extensively for terrestrial plant/bacterial systems, but sulfation is not present in these cases, meaning the marine enzymes used to degrade carrageenans must possess unique features to recognize these modifications. To gain insights into these features, we have focused on κ-carrageenases from two distant bacterial phyla, which belong to glycoside hydrolase family 16 (GH16) and cleave the β-1,4 linkage of κ-carrageenan. We have solved the crystal structure of the catalytic module of ZgCgkA from Zobellia galactanivorans at 1.66 Å resolution, and compared it with the only other structure available, that of PcCgkA from Pseudoalteromonas carrageenovora 9T (ATCC 43555T). We also describe the first substrate complex in the inactivated mutant form of PcCgkA at 1.7 Å resolution. The structural and biochemical comparison of these enzymes suggests key determinants that underlie the functional properties of this subfamily. In particular, we identify several arginine residues that interact with the polyanionic substrate, and confirmed the functional relevance of these amino acids using a targeted mutagenesis strategy. These results give new insight in the diversity of the κ-carrageenase subfamily. The phylogenetic analyses show the presence of several distinct clades of enzymes that relate to differences in modes of action or subtle differences within the same substrate specificity, matching the hybrid character of the κ-carrageenan polymer.

Publisher URL: http://doi.org/10.1074/jbc.M117.808279

DOI: 10.1074/jbc.M117.808279