ι-Carrageenases constitute a novel family of glycoside hydrolases, unrelated to that of κ-carrageenases

i-Carrageenases are polysaccharide hydrolases that cleave the beta -1,4 linkages between the D-galactose-4-sulfate and 3,6-anhydro-D-galactose-2-sulfate residues in the red algal galactans known as i-carrageenans. We report here on the purification of i-carrageenase activity from the marine bacterium Zobeellia galactanovorans and on the characterization of L-carrageenase structural. genes. Genomic libraries from this latter bacterium as well as from Alteromonas fortis were functionally screened for the presence of i-carrageenase(+) clones. The Z. galactanovorans and a fortis L-carrageenase genes encode homologous proteins of 53.4 and 54.8 kDa, respectively. Based on hydrophobic cluster analysis and on the H-1 NMR monitoring of the products of the overexpressed A. fortis L-carrageenase, these enzymes appear to form a new family of glycoside hydrolases, unrelated to that of kappa -carrageenases and with an inverting mechanism of hydrolysis. They both feature a 45-amino acid-long N-terminal segment with sequence similarity to the N-terminal region of several other polysaccharidases, In those for which a three-dimensional structure is available, this conspicuous segment, also deemed glycanase motif (Chua, J, E, H,, Manning, P, A., and Morons, R, (1999) Microbiology (Reading) 145, 1649-1659), corresponds to a strand-helix-strand cap that covers the N-terminal end of a common, right-handed beta -helical fold.