Structure and organization of the hisA gene of the thermophilic archaebacterium Methanococcus thermolithotrophicus.

A restriction fragment of Methanococcus thermolithotrophicus genomic DNA was cloned into pUC8 to produce plasmid pET9301, which complements mutations in the hisA gene of Escherichia coli. Sequencing the DNA (2,155 base pairs) cloned from this thermophilic methanogen demonstrated that the M. thermolithotrophicus hisA gene is located within a cluster of open reading frames (ORFs) and is 68 and 69% homologous at the nucleotide level to the hisA genes of the mesophilic methanococci M. voltae and M. vannielii, respectively. The ORF (ORF 206) immediately 5' to the hisA gene of M. thermolithotrophicus is partially deleted in the genomes of the two mesophilic species, whereas ORF 114, which is 5' to ORF 206, is conserved in all three species.     

Conservation of structure in the human gene encoding argininosuccinate synthetase and the argG genes of the archaebacteria Methanosarcina barkeri MS and Methanococcus vannielii.

The DNA sequences of the argG genes of Methanosarcina barkeri MS and Methanococcus vannielii were determined. The polypeptide products of these methanogen genes have amino acid sequences which are 50% identical to each other and 38% identical to the amino acid sequence encoded by the exons of the human argininosuccinate synthetase gene. Introns in the human chromosomal gene separate regions which encode amino acids conserved in both the archaebacterial and human gene products. An open reading frame immediately upstream of argG in Methanosarcina barkeri MS codes for an amino acid sequence which is 45 and 31% identical to the sequences of the large subunits of carbamyl phosphate synthetase in Escherichia coli and Saccharomyces cerevisiae, respectively. If this gene encodes carbamyl phosphate synthetase in Methanosarcina barkeri, this is the first example, in an archaebacterium, of physical linkage of genes that encode enzymes which catalyze reactions in the same amino acid biosynthetic pathway.     

Autogenous translational regulation of the ribosomal MvaL1 operon in the archaebacterium Methanococcus vannielii.

The mechanisms for regulation of ribosomal gene expression have been characterized in eukaryotes and eubacteria, but not yet in archaebacteria. We have studied the regulation of the synthesis of ribosomal proteins MvaL1, MvaL10, and MvaL12, encoded by the MvaL1 operon of Methanococcus vannielii, a methanogenic archaebacterium. MvaL1, the homolog of the regulatory protein L1 encoded by the L11 operon of Escherichia coli, was shown to be an autoregulator of the MvaL1 operon. As in E. coli, regulation takes place at the level of translation. The target site for repression by MvaL1 was localized by site-directed mutagenesis to a region within the coding sequence of the MvaL1 gene commencing about 30 bases downstream of the ATG initiation codon. The MvaL1 binding site on the mRNA exhibits similarity in both primary sequence and secondary structure to the L1 regulatory target site of E. coli and to the putative binding site for MvaL1 on the 23S rRNA. In contrast to other regulatory systems, the putative MvaL1 binding site is located in a sequence of the mRNA which is not in direct contact with the ribosome as part of the initiation complex. Furthermore, the untranslated leader sequence is not involved in the regulation. Therefore, we suggest that a novel mechanism of translational feedback regulation exists in M. vannielii.     

Sequence of three copies of the gene for the major Drosophila heat shock induced protein and their flanking regions

The primary sequence of the major heat shock gene of D. melanogaster, that for the 70,000 protein, has been determined. One of the reading frames is devoid of stop codons for over 2000 bp. The region between the first ATG and the first stop codon encodes a protein of molecular weight 70,270. The 5' end of the messenger RNA was localized in the DNA sequence by two independent methods. The 5' flanking sequences of three distinct 70K genes were also determined. Extensive homology in the primary sequences extends about 500 bp upstream from the ATG, which is the presumptive initiation of protein synthesis. Each 70K gene has the putative promoter sequence TATAAATA about 325 bp upstream from this ATG. A heptanucleotide sequence identified as the capping site for other messengers is found 24-30 bp downstream from the ends of the A-T-rich sequence. A 12 bp sequence with dyad symmetry begins 23 bp upstream from the beginning of the above A-T-rich sequence.