Cloning, sequencing and expression of the Taq I restriction-modification system.
The Taq I modification and restriction genes (recognition sequence TCGA) have been cloned in E. coli and their DNA sequences have been determined. Both proteins were characterized and the N-terminal sequence of the endonuclease was determined. The genes have the same transcriptional orientation with the methylase gene 5' to the endonuclease gene. The methylase gene is 1089 bp in length (363 amino acids, 40,576 daltons); the endonuclease gene is 702 bp in length (234 amino acids, 27,523 daltons); they are separated by 132 bp. Both methylase and endonuclease activity can be detected in cell extracts. The clones fully modify the vector and chromosomal DNA but they fail to restrict infecting phage. Clones carrying only the restriction gene are viable even in the absence of modification. The restriction gene contains 7 Taq I sites; the modification gene contains none. This asymmetric distribution of sites could be important in the regulation of the expression of the endonuclease gene.
Slatko
B E
BE
New England Biolabs, Inc., Beverly, MA 01915.
Benner
J S
JS
Jager-Quinton
T
T
Moran
L S
LS
Simcox
T G
TG
Van Cott
E M
EM
Wilson
G G
GG
D000595:Amino Acid Sequence; D010582:Bacteriophage lambda / Q000235:genetics; D001483:Base Sequence; D002874:Chromosome Mapping; D003001:Cloning, Molecular / Q000379:methods; D003062:Codon; D004262:DNA Restriction Enzymes / Q000235:genetics* / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000235:genetics* / Q000302:isolation & purification; D015252:Deoxyribonucleases, Type II Site-Specific; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D013045:Species Specificity; D013824:Thermus / Q000201:enzymology / Q000235:genetics
D003062:Codon; D004269:DNA, Bacterial; D004262:DNA Restriction Enzymes; D015252:Deoxyribonucleases, Type II Site-Specific; C027046:TCGA-specific type II deoxyribonucleases