Multiple endoglycosidase (Endo) F activities expressed by Flavobacterium meningosepticum. Endo F1: molecular cloning, primary sequence, and structural relationship to Endo H.
A full-length insert for the endo-beta-N-acetylglucosaminidase (Endo) F1 gene was located on a 2,200-base pair EcoRI fragment of genomic DNA and cloned into the plasmid vector Bluescript. Transformed Escherichia coli cells expressed Endo F1 activity very well, but the enzyme apparently was not processed and secreted into the medium as it normally is in Flavobacterium meningosepticum. DNA sequencing revealed an open reading frame of 1,017 nucleotides encoding a putative 50-amino acid signal sequence, and a mature protein (31,667 Da) of 289 amino acids. The deduced amino acid sequence was verified by direct Edman microsequencing of 88% of the purified protein as tryptic and V8 protease peptides. Alignment of Endo F1 (289 amino acids) with the established amino acid sequence of Streptomyces plicatus Endo H (271 amino acids) revealed a 32% structural identity over the entire sequence and a high degree of conservative replacements. Potential catalytic domains identified in other proteins that hydrolyze the beta 1,4 glycosidic linkage between N-acetylglucosamine residues are also conserved for amino acid identity and relative spacing in Endo F1.
Tarentino
A L
AL
Division of Clinical Sciences, Wadsworth Center for Laboratories and Research, New York State Department of Health, Albany 12201-0509.
Quinones
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G
Schrader
W P
WP
Changchien
L M
LM
Plummer
T H
TH
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001483:Base Sequence; D002384:Catalysis; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D005417:Flavobacterium / Q000201:enzymology*; D006596:Hexosaminidases / Q000235:genetics* / Q000378:metabolism; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010957:Plasmids; D016415:Sequence Alignment; D013302:Streptomyces / Q000201:enzymology
D000596:Amino Acids; D004269:DNA, Bacterial; D006596:Hexosaminidases; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase