Files in this item



application/pdf9136702.pdf (8MB)Restricted to U of Illinois
(no description provided)PDF


Title:Cloning, sequencing, and biochemical characterization of endo-N-acylneuraminidase
Author(s):Petter, Jean Guard
Doctoral Committee Chair(s):Vimr, Eric R.
Department / Program:Veterinary Biosciences
Discipline:Veterinary Medical Science
Degree Granting Institution:University of Illinois at Urbana-Champaign
Subject(s):Biology, Molecular
Biology, Neuroscience
Chemistry, Biochemistry
Abstract:Bacteriophage K1F Endo-N-acylneuraminidase hydrolyzes alpha-2,8-linked polysialic acid found on K1 encapsulated Escherichia coli and on the neural cell adhesion molecule (NCAM). A novel cloning strategy, which pursued ligation of partially restricted ALU 1 or SAU 3A genomic K1F DNA fragments, is described. Sequence analysis reveals that Endo-N is a domainally arranged, 920 amino acid protein with a T7-like gene organization. A 200 amino acid N-terminal domain is homologous to the T7 tail protein N-terminus (gene 17). A 520 amino acid middle domain is possibly similar to the Sea Urchin arylsulfatase precursor, as determined by statistical processing of jumbled sequences. Results from comparative analyses of epitope and nucleotide similarities to PK1E endoneuraminidase indicate that a 200 amino acid C-terminal domain, unique to K1F Endo-N, could be responsible for stabilizing K1F Endo-N to proteolysis. In addition to sequence analysis, biochemical characterization indicates that the molecule is a dimer of identical 102 kDa polypeptide chains. Immunoblot analysis of SDS-PAGE fractionated Endo-N indicates that no post-transcriptional modifications are necessary for expression of native protein. A method for detecting molecular weight variants, which could be indicative of in vivo protein folding intermediates as found in the P22 tail spike protein, is presented. Future research with Endo-N depends on developing an expression vector. Biochemical and nucleotide sequence results presented herein indicate that construction of an Endo-N expression vector is possible.
Issue Date:1991
Rights Information:Copyright 1991 Petter, Jean Guard
Date Available in IDEALS:2011-05-07
Identifier in Online Catalog:AAI9136702
OCLC Identifier:(UMI)AAI9136702

This item appears in the following Collection(s)

Item Statistics