Nucleotide sequence of the phoS gene, the structural gene for the phosphate-binding protein of Escherichia coli.

phoS is the structural gene for the phosphate-binding protein, which is localized in periplasm and involved in active transport of phosphate in Escherichia coli. It is also a negative regulatory gene for the pho regulon, and the gene expression is inducible by phosphate starvation. The complete nucleotide sequence of the phoS gene was determined by the method of Maxam and Gilbert (A. M. Maxam and W. Gilbert, Methods Enzymol. 65:499-560, 1980). The amino acid sequences at the amino termini of the pre-PhoS and PhoS proteins and at the carboxy terminus of the PhoS protein were determined by using the purified proteins. Furthermore, the amino acid sequence of enzymatically digested peptide fragments of the PhoS protein was determined. The combined data established the nucleotide sequence of the coding region and the amino acid sequence of the pre-PhoS and the PhoS proteins. The pre-PhoS protein contains an extension of peptide composed of 25 amino acid residues at the amino terminus of the PhoS protein, which has the general characteristics of a signal peptide. The mature PhoS protein is composed of 321 amino acid residues, with a calculated molecular weight of 34,422, and lacks the disulfide bond and methionine. The regulatory region of phoS contains a characteristic Shine-Dalgarno sequence at an appropriate position preceding the translational initiation site, as well as three possible Pribnow boxes and one -35 sequence. the nucleotide sequence of the regulatory region of phoS was compared with those of phoA and phoE, the genes constituting the pho regulon.     

Intragenic suppression in the uvrD gene of Escherichia coli. I. Temperature-sensitive uvrD mutations

A temperature-sensitive uvrD mutant, HD323 uvrD4, was isolated from the uvrD mutant HD4 uvrD3. The temperature sensitivity of the uvrD4 gene product was reversible. The suppressor mutation uvrD44 which rendered the uvrD3 mutant temperature-sensitive could be separated from the uvrD3 mutation by replacing the PstI fragment, which encodes the C-terminal half of the UvrD protein. The uvrD44 mutation was found to make host bacteria lethal at non-permissive temperatures only when cloned on a low copy vector pMF3. The nucleotide sequence of the uvrD3 and uvrD4 mutant genes was determined. The nucleotide change found in the uvrD3 at +1235, GAA to AAA, only alters the amino acid sequence from Glu at 387 to Lys. The uvrD44 has another nucleotide change at +1859, GAA to AAA (Glu at 595 to Lys), which is considered to be the suppressor mutation uvrD44.     

The nucleotide sequence of the uvrD gene of E. coli.

The nucleotide sequence of a cloned section of the E. coli chromosome containing the uvrD gene has been determined. The coding region for the UvrD protein consists of 2,160 nucleotides which would direct the synthesis of a polypeptide 720 amino acids long with a calculated molecular weight of 82 kd. The predicted amino acid sequence of the UvrD protein has been compared with the amino acid sequences of other known adenine nucleotide binding proteins and a common sequence has been identified, thought to contribute towards adenine nucleotide binding.     

Nucleotide sequence studies of polyoma DNA. The Hpa II 3/5 junction to the Hpa II 4/Hae III 18 junction, encoding the origin of DNA replication and the 5' end of the early region.

The nucleotide sequence of polyoma DNA, from near the Hpa II 3/5 unction to the Hpa II 4/ae III 18 junction has been determined by the chemical method of Maxam and Gilbert (Maxam, A., and Gilbert, W. (1977) Proc. Natl. Acad. Sci. U. S. A. 74, 560--564). The sequence contains 878 base paris, including the origin of DNA replication and the region known to encode the hr-t function. The region corresponding to the origin of DNA replication contains several short-repeated sequences and palindromes. There is a 30-base-pair region with striking similarity to the corresponding region of SV40, and, as in SV40, a portion of that sequence is capable of forming a stable hairpin loop. In the region encoding the hr-t function, there is apparently a single open reading frame extending from position 188 to theHpa III 4/Hae III 18 junction. The potential translation product of this open frame begins with an initiation codon starting at position 188, and the first five amino acids of this product are Met-Asp-Arg-Val-Leu. This sequence is similar to the NH2-terminal five amino acids of SV40 small t-antigen known from nucleotide and amino acid sequencing to be Met-Asp-Lys-Val-Leu.     

Cloning and sequence of the crp gene of Escherichia coli K 12.

We have determined the nucleotide sequence of the crp gene of Escherichia coli K 12. From a lambda transducing phage, the crp region was subcloned into pBR322. The gene was localized on the cloned fragment by determining the length of deletions which affect its expression. Its nucleotide sequence was established by using the technique of Maxam and Gilbert. The deduced amino-acid sequence is in agreement with the previously published amino acid composition of the protein (1, 2). Analysis of the sequence confirms that the DNA binding domain is located in the C-terminal portion of the protein.     

Nucleotide sequence of the restriction fragment Hind-F-EcoRI1 of simian-virus-40 DNA (part of the VP1 gene).

The nucleotide sequence of the simian virus 40 (SV40) genome region between the cleavage sites for restriction endonucleases EcoRI (map position 0) and HindII (map position 0.05) has been determined mainly by the partial chemical DNA degradation procedure of Maxam and Gilbert. This fragment represents 5.3% of the genome of SV40 and is located in the late region, internally in the VP1 gene. The message strand shows only one open reading frame for translation into protein, which connects to the one for the preceding fragment. On this basis part of the amino acid sequence of the VP1 protein is presented.     

Overlapping of the VP2-VP3 gene and the VP1 gene in the SV40 genome.

The nucleotide sequence of the SV40 Hind E fragment has been determined mainly by the partial chemical degradation procedure of Maxam and Gilbert (1977). The sequence of the strand with the same polarity as the late messenger RNA shows only one open reading frame for translation. Considering that VP3 corresponds to the carbosyl terminal part of VP2, and considering various evidence which indicates that the SV40 Hind E segment is part of the amino acid sequence of VP2-VP3. It continues clockwise in Hind K, where it terminates with a UAA signal. The latter is located 110 nucleotides beyond the initiation signal for the major structural protein VP1 (Fiers et al., 1975; Van de Voorde et al., 1976). Hence this small overlapping region of the genome codes for the synthesis of three different proteins in two different reading frames. The deduced amino acid sequence covers a major part of the vp3 poly peptide, and the amino acid composition is in good agreement with published values (Greenaway and Levine, 1973).     

DNA sequence of the araC regulatory gene from Escherichia coli B/r.

The DNA sequence of the araC regulatory gene from Escherichia coli B/r has been determined by the base-specific chemical cleavage reactions of Maxam and Gilbert. An open reading frame is found which codes for a protein of 292 amino acids. A nonsense mutation, araC5, is shown to result from a G to A transition at nucleotide 429 converting the tryptophan codon TGG to the amber codon TAG. A deletion which does not recombine with any known point mutation in araC, delta(araCO)719, removes all but the last 22 codons of the gene.