Cloning, sequencing, and enhanced expression of the dihydropteroate synthase gene of Escherichia coli MC4100.
The Escherichia coli gene coding for dihydropteroate synthase (DHPS) has been cloned and sequenced. The protein has 282 amino acids and a compositional molecular mass of 30,314 daltons. Increased expression of the enzyme was realized by using a T7 expression system. The enzyme was purified and crystallized. A temperature-sensitive mutant was isolated and found to express a DHPS with a lower specific activity and lower affinities for para-aminobenzoic acid and sulfathiazole. The allele had a point mutation that changed a phenylalanine codon to a leucine codon, and the mutation was in a codon that is conserved among published DHPS sequences.
Dallas
W S
WS
Department of Molecular Genetics and Microbiology, Burroughs Wellcome Co., Research Triangle Park, North Carolina 27709.
Gowen
J E
JE
Ray
P H
PH
Cox
M J
MJ
Dev
I K
IK
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D004094:Dihydropteroate Synthase / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D015723:Gene Library; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009690:Nucleic Acid Conformation; D012333:RNA, Messenger; D011994:Recombinant Proteins / Q000096:biosynthesis*; D012689:Sequence Homology, Nucleic Acid
D012333:RNA, Messenger; D011994:Recombinant Proteins; D004094:Dihydropteroate Synthase