Identification of an EF-Tu protein that is periplasm-associated and processed in Neisseria gonorrhoeae.
A 44 kDa protein is a dominant component of periplasmic extracts of Neisseria gonorrhoeae. Peptide sequence generated from a cyanogen-bromide-cleaved fragment of this protein indicated sequence homology with elongation factor-Tu (EF-Tu). Polyclonal antiserum was made against the 44 kDa protein purified from periplasm extracts of N. gonorrhoeae. The preabsorbed antiserum was immunoblotted against whole-cell lysates on two-dimensional gels. A 44 kDa protein and a smaller 37 kDa protein were recognized by this antiserum. A N. gonorrhoeae gamma phage DNA library was screened and a clone expressing a 44 kDa protein was identified. The DNA insert in this clone contained several genes homologous to genes contained in the str operon of Escherichia coli. One ORF product with a calculated molecular mass of 43 kDa was highly homologous to the EF-TuA of E. coli. A synthetic peptide antiserum specific for a portion of the C terminus of EF-Tu confirmed that the 37 kDa protein in whole-cell lysates of N. gonorrhoeae was a processed form of EF-Tu. Deletion of the tufA gene homologue in N. gonorrhoeae was attempted but was unsuccessful.
Porcella
S F
SF
Department of Microbiology, University of Texas Southwestern Medical School at Dallas 75235, USA. porcella@utsw.swmed.edu
Belland
R J
RJ
Judd
R C
RC
D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D015180:Electrophoresis, Gel, Two-Dimensional; D004926:Escherichia coli / Q000235:genetics; D015723:Gene Library; D015151:Immunoblotting; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009344:Neisseria gonorrhoeae / Q000235:genetics* / Q000276:immunology; D010444:Peptide Elongation Factor Tu / Q000235:genetics* / Q000276:immunology; D010449:Peptide Mapping; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D016133:Polymerase Chain Reaction; D016364:Reading Frames; D017384:Sequence Deletion
D010455:Peptides; D010444:Peptide Elongation Factor Tu