Independent genes coding for three acidic proteins of the large ribosomal subunit from Saccharomyces cerevisiae

The yeast ribosome contains three acidic proteins, L44, L44', and L45, closely related from a structural point of view, that seem to play a functional role similar to that of proteins L7 and L12 in the bacterial ribosome. By screening a cDNA bank in lambda gt11 with specific polyclonal and monoclonal antibodies, recombinant phages expressing each one of the acidic proteins have been cloned. A unique copy of each gene is detected using the phage cDNA inserts as probes in nitrocellulose blots of yeast DNA digested with different restriction enzymes. The inserts were subcloned in the plasmid pUC19, and their physical maps and nucleotide sequences were determined. By using the cDNA inserts as probes in genomic DNA banks, DNA fragments carrying the acidic protein genes have been cloned, characterized, and sequenced. The results conclusively show that the three yeast acidic proteins are coded by independent genes and are not the result of a post-translational modification of the product of a unique gene, as in bacteria. Like most ribosomal protein genes, the gene for protein L44' has an intron and two upstream stimulatory boxes (UASrpg) fitting closely to the consensus sequence. The genes coding for proteins L44 and L45 lack introns and seem also exceptional in other characteristics of their sequences. Proteins L44 and L45 have amino acid sequences with about 80% similarity. Protein L44' is only 63% similar to the other two polypeptides. The three proteins have highly conserved carboxyl termini comprising the last 30 amino acids, and the first 10 amino acids of L44 and L45 are identical. The results cast doubts about the possibility of a similar role for the different acidic ribosomal proteins.     

Structural comparison of yeast ribosomal protein genes.

The primary structure of the genes encoding the yeast ribosomal proteins L17a and L25 was determined, as well as the positions of the 5'- and 3'-termini of the corresponding mRNAs. Comparison of the gene sequences to those obtained for various other yeast ribosomal protein genes revealed several similarities. In all split genes the intron is located near the 5'-side of the amino acid coding region. Among the introns a clear pattern of sequence conservation can be observed. In particular the intron-exon boundaries and a region close to the 3'-splice site show sequence homology. Conserved sequences were also found in the leader and trailer regions of the ribosomal protein mRNAs. The 5'-flanking regions of the yeast ribosomal protein genes appeared to contain sequence elements that many but not all ribosomal protein genes have in common, and therefore may be implicated in the coordinate expression of these genes. The amino acid coding sequences of the ribosomal protein genes show a biased codon usage. Like most yeast ribosomal protein molecules, L17a and L25 are particularly basic at their N-terminus.     

Genomic DNA sequence for human C-reactive protein

The gene for the prototype acute phase reactant, C-reactive protein, has been isolated from two lambda phage libraries containing inserted human DNA fragments using synthetic oligonucleotide probes. Nucleotide sequence analysis indicates that after coding for a signal peptide of 18 amino acids and the first two amino acids of the mature protein, there is an intron of 278 base pairs followed by the nucleotide sequence for the remaining 204 amino acids. The intron is unusual in that it contains on the positive strand a poly(A) stretch 16 nucleotides long and a poly(GT) region 30 nucleotides long which could adopt the Z-form of DNA. The nucleotide sequence reported here confirms the amino acid sequence of mature C-reactive protein as originally reported except that it codes for an additional 19 amino acids beginning at position 62. Thus DNA sequence analysis predicts that the mature protein consists of 206 amino acids rather than 187 as originally reported. The mRNA cap site is located 104 nucleotides from the start of the signal peptide and there is a 3' noncoding region 1.2 kilobase pairs in length. The gene has a typical promoter containing the sequences TATAAAT and CAAT 29 and 81 base pairs upstream, respectively, of the cap site.     

Structure of the mouse C-reactive protein gene

A genomic DNA clone corresponding to the mouse C-reactive protein (CRP) has been isolated and characterized. The mouse CRP gene is 1.9-kilobase pairs in length and contains a single intron of 213-base pairs which interrupts the codon for the 2nd amino acid residue of the mature CRP protein. We compared nucleotide sequences of the mouse and human CRP genes and discussed structures of possible regulatory sequences. With this characterization, the isolation and sequence analyses of a set of mouse and human pentraxin genes, i.e. CRP and serum amyloid P component genes is not complete.     

Complete nucleotide sequence of recD, the structural gene for the alpha subunit of Exonuclease V of Escherichia coli.

Intracellular amplification of the Escherichia coli RecB and RecC proteins does not result in an increase in Exonuclease V activity unless the level of a third protein, encoded between the recB and argA genes, is also amplified. Nucleotide sequence analysis of this region reveals a 1,824 nucleotide open reading frame which would encode a protein of 608 amino acids with a calculated molecular weight of 66,973. This is assumed to be the structural gene for the alpha subunit of Exonuclease V, recently designated recD. The proposed initiation codon of the recD gene overlaps the termination codon of the upstream recB gene by one nucleotide, suggesting that these genes may form an operon. The deduced amino acid sequence of the RecD protein contains a region which is homologous to highly conserved sequences in adenine nucleotide binding proteins.     

The nucleotide sequence of the yeast MEL1 gene.

The complete nucleotide sequence of the MEL1 gene of the yeast, Saccharomyces cerevisiae, encoding alpha-galactosidase was determined. The nucleotide sequence contains an open reading frame of 1413 bp encoding a protein of 471 amino acids. Comparison with the known N-terminal amino acid sequence of the mature secreted protein indicated that alpha-galactosidase is synthesized as a precursor with an N-terminal signal sequence of 18 amino acids. The general features of this signal peptide resemble those of other yeast signal peptides. Molecular weight of the mature alpha-galactosidase polypeptide deduced from the nucleotide sequence is 50.049 kd. The 5' regulatory region has sequences in common with other yeast genes regulated by the GAL4-protein.     

Nucleotide sequences of tobacco chloroplast genes for elongator tRNAMet and tRNAVal (UAC): the tRNAVal (UAC) gene contains a long intron.

The nucleotide sequences of tobacco chloroplast genes for elongator tRNAMet and tRNAVal (UAC) have been determined. The tRNAVal gene contains a 571 base pairs intron located in the anticodon loop. The tRNAVal gene is transcribed as a 750 bases precursor RNA molecule. Both tRNAs deduced from the DNA sequences show 97% sequence homologies with those of spinach chloroplasts.     

Nucleotide sequence and intron structure of the apocytochrome b gene of Neurospora crassa mitochondria

The sequence of the apocytochrome b (cob) gene of Neurospora crassa has been determined. The structural gene is interrupted by two intervening sequences of approximately 1260 bp each. The polypeptide encoded by the exons shows extensive homology with the cob proteins of Aspergillus nidulans and Saccharomyces cerevisiae (79% and 60%, respectively). The two introns are, however, located at sites different from those of introns in the cob genes of A. nidulans and S. cerevisiae (which contain highly homologous introns at the same site within the gene). The introns share several short regions of sequence homology (10-12 bp long) with each other and with other fungal mitochondrial introns. Moreover, the second intron contains a 50 nucleotide long sequence that is highly homologous with sequences within every ribosomal intron of fungal mitochondria sequenced to date. The conserved sequences may allow the formation of a core secondary structure, which is nearly identical in many mitochondrial introns. The conserved secondary structure may be required for intron splicing. The second intron contains an open reading frame, continuous with the preceding exon, of approximately 290 codons. Two stretches of 10 amino acid residues, conserved in many introns, are present in the open reading frame.     

Nucleotide sequence of the O gene and of the origin of replication in bacteriophage lambda DNA.

The nucleotide sequence of the O gene in bacteriophage lambda DNA is presented. According to two possible initiator codons, the primary structure of the O protein deduced from the DNA sequence consists of 278 or 299 amino acid residues. Structure and function of the O protein--one of the two phage initiator proteins for lambda DNA replication--are discussed in the light of a secondary structure model for the O protein. The central part of the O gene contains a cluster of symmetrical sequences extending over 160 base pairs. The point mutation of the cis-dominant replication mutant ti12 is located in this region.     

Yeast ribosomal proteins: XII. YS11 of Saccharomyces cerevisiae is a homologue to E. coli S4 according to the gene analysis.

We isolated and sequenced a gene, YS11A, encoding ribosomal protein YS11 of Saccharomyces cerevisiae. YS11A is one of two functional copies of the YS11 gene, located on chromosome XVI and transcribed in a lower amount than the other copy which is located on chromosome II. The disruption of YS11A has no effect on the growth of yeast. The 5'-flanking region contains a similar sequence to consensus UASrpg and the T-rich region. The open reading frame is interrupted with an intron located near the 5'-end. The predicted amino acid sequence reveals that yeast YS11 is a homologue to E. coli S4, one of the ram proteins, three chloroplast S4s and others out of the ribosomal protein sequences currently available.     

Nucleotide sequence and DNA recognition elements of alc, the structural gene which encodes allantoicase, a purine catabolic enzyme of Neurospora crassa

The nitrogen regulatory circuit of Neurospora crassa contains structural genes that encode nitrogen catabolic enzymes which are subject to complex genetic and metabolic regulation. This set of genes is controlled by nitrogen limitation, by specific induction, and by the action of nit-2, a major positive-acting regulatory gene, and nmr, a negative-acting control gene. The complete nucleotide sequence of alc, the gene that encodes allantoicase, a purine catabolic enzyme, is presented. The alc gene contains a single intron, is transcribed from two initiation sites situated approximately 50 nb upstream of the translation start site, and encodes a protein comprised of 354 amino acids. Mobility shift and DNA footprint experiments identified a single binding site for the NIT2 regulatory protein in the alc promoter region. The binding site contains a 10 nucleotide base pair symmetrical sequence which is flanked by two possible core binding sequences, TATCT and TATCG. Mutant NIT2/beta-gal fusion proteins with amino acid substitutions in a putative zinc-finger motif were shown to be completely deficient in the ability to bind to the alc promoter DNA fragment.