Isolation and sequence analysis of a cDNA encoding rat liver alpha-L-fucosidase.
cDNA clones for alpha-L-fucosidase were isolated from a rat liver lambda gt11 expression library by using both monospecific polyclonal antibodies against the affinity-purified enzyme and biotinylated rat liver fucosidase cDNA sequences as probes. The largest clone, lambda FC9, contained a 1522 bp full-length cDNA insert (FC9) that encoded the 434-amino acid-residue subunit (Mr 50439) of rat liver alpha-L-fucosidase. A putative signal peptide 28 amino acid residues in length preceded the sequence for the mature protein. In addition, FC9 specified for 11 nucleotide residues of 5' untranslated sequence, 78 nucleotide residues of 3' untranslated sequence and a poly(A) tail. The deduced amino acid sequence from FC9 in conjunction with the experimentally determined N-terminus of the mature enzyme suggested that rat liver fucosidase did not contain a pro-segment. However, there was the possibility of limited N-terminal processing (one to five amino acid residues) having occurred after removal of the predicted signal peptide. Amino acid sequences deduced from FC9 were co-linear with amino acid sequences measured at the N-terminus of purified fucosidase and on two of its CNBr-cleavage peptides. An unusual aspect of rat liver alpha-L-fucosidase protein structure obtained from the FC9 data was its high content of tryptophan (6%). The coding sequence from FC9 showed 82% sequence identity with that from a previously reported incomplete human fucosidase sequence [O'Brien, Willems, Fukushima, de Wet, Darby, DiCioccio, Fowler & Shows, (1987) Enzyme 38, 45-53].
Fisher
K J
KJ
Department of Molecular and Cell Biology, Althouse Laboratory, Pennsylvania State University, University Park 16802.
Aronson
N N
NN
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics / Q000302:isolation & purification*; D004023:Dictyostelium / Q000201:enzymology / Q000235:genetics; D000939:Epitopes / Q000032:analysis; D004926:Escherichia coli / Q000235:genetics; D015723:Gene Library; D005796:Genes; D008099:Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000302:isolation & purification; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D005644:alpha-L-Fucosidase / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification
D000939:Epitopes; D011993:Recombinant Fusion Proteins; D004247:DNA; D005644:alpha-L-Fucosidase