Cathepsin, a major protease of the marine sponge Geodia cydonium: purification of the enzyme and molecular cloning of cDNA.
Sponges are suspension-feeders that are devoid of body cavities. Phagocytosis is the major route of nutrition in these animals. In an attempt to understand protein digestion, cathepsin was identified in crude extracts from the sponge Geodia cydonium. This enzyme was purified from lysosomes by a two-step procedure--pH precipitation and FPLC separation--to apparent homogeneity; it showed an M(r) of 26,000. Inhibitor as well as substrate studies showed that the sponge cathepsin belongs to the subfamily L of these cysteine proteases. The complete cDNA coding for cathepsin L was isolated and characterized. The deduced aa sequence contains 322 residues, has an M(r) of 36,085, and shows the characteristic signatures known from other cathepsins of the L subfamily: e.g., cleavage site for the proregion, the ERFNIN motif, and the conserved regions forming the catalytic triad of cysteine proteases. Phylogenetic analyses revealed that the sponge sequence groups with the cathepsin L subfamily and branches off first from the other metazoan members. The sponge sequence shows high homology to that isolated from Dictyostelium discoideum and only low similarity to the protozoan cathepsins L from Paramecium tetraurelia and Tetrahymena thermophila. From the data presented it is concluded that cathepsin L is the major digestive protease in sponges.
Krasko
A
A
Abteilung Angewandte Molekularbiologie, Universität, Duesbergweg, Mainz, Germany.
Gamulin
V
V
Seack
J
J
Steffen
R
R
Schröder
H C
HC
Müller
W E
WE
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002403:Cathepsins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D018076:DNA, Complementary; D006801:Humans; D007700:Kinetics; D008247:Lysosomes / Q000201:enzymology; D008969:Molecular Sequence Data; D010802:Phylogeny; D011161:Porifera / Q000145:classification / Q000201:enzymology* / Q000235:genetics*; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D011994:Recombinant Proteins; D002403:Cathepsins