Isolation and characterization of a fructosyltransferase gene from Streptococcus mutans GS-5.
AUTOR(ES)
Sato, S
RESUMO
A fructosyltransferase (FTF) gene from Streptococcus mutans GS-5 has been isolated from a lambda L47.1 clone bank. The gene was contained on an 11.7-kilobase GS-5 DNA fragment and was initially subcloned into plasmid pACYC184 as a 5.4-kilobase HindIII fragment. However, further analysis revealed that transcription of the FTF gene was initiated at the P1 promoter contained on the vector. It was possible to subclone the FTF gene with its presumed promoter as a 3.4-kilobase EcoRI fragment to produce the chimeric plasmid pSS22 expressing FTF activity. The cloned enzyme was purified to apparent homogeneity after ammonium sulfate precipitation, gel filtration, and DEAE-Bio-Gel-A chromatography followed by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The purified enzyme displayed a lower molecular weight (63,000) compared with the multiple activities detected in the culture fluids of strain GS-5. In addition, storage of the purified enzyme resulted in the formation of even lower-molecular-weight enzymatically active species. These results suggested that proteolytic degradation of the FTF occurs both in S. mutans and in Escherichia coli. In addition, a comparison of the properties of the cloned enzyme with those previously characterized from another serotype c S. mutans strain suggests that multiple FTF genes may be present in these organisms.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=262214Documentos Relacionados
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