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 proce...

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Bibliographic Details
Published in:The Journal of biological chemistry 1992-02, Vol.267 (6), p.3868-3872
Main Authors: A L Tarentino, G Quinones, W P Schrader, L M Changchien, T H Plummer, Jr
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Amino Acids - analysis
Base Sequence
Catalysis
Cloning, Molecular
DNA, Bacterial - genetics
Flavobacterium - enzymology
Hexosaminidases - genetics
Hexosaminidases - metabolism
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase - genetics
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase - metabolism
Molecular Sequence Data
Plasmids
Sequence Alignment
Streptomyces - enzymology
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Summary: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.
ISSN:0021-9258
1083-351X
DOI:10.1016/s0021-9258(19)50606-8