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Abstract We have introduced a consistent nomenclature for the various subunits of the NatA-NatE N-terminal acetyltransferases from yeast, humans and other eukaryotes.

The BTN1 gene product of Saccharomyces cerevisiae bears 39% identity and 59% similarity to human CLN3 protein associated with the Batten disease. The deletion of the associated genes lowers the buffering capacity of the cytoplasm leading to disturbances in the intracellular pH homeostasis, which is linked to the disease manifestation.

Modification of ribosomes by a variant multiubiquitin chain and regulated by the cell cycle is discussed. The regulatory role of the muliubiquitin chains seems to be reversible and not to target the acceptor protein for degradation.The substrate found to be in question is now identified as ribosomal protein L28.

The pH-dependent inhibition of 22 metal salts was systematically examined for the yeast Saccharomyces cerevisiae. It was observed that the inhibition of growth by Cu, Co or Ni salts is substantially increased by histidine auxotrophy and by increasing the pH of the medium. Deletion of the high-affinity histidine permease Hip1p in His- strains caused greater sensitivity to Cu, Co and Ni and the requirement of a higher level of histidine to reverse the inhibition. It was concluded that intracellular histidine reduces the pH-dependent toxicity of Cu, Co and Ni.

The Cct double-ring chaperonin complex of Saccharomyces cerevisiae is comprised of eight essential subunits, Cct1p-Cct8p, and assists the folding of substrates such as actins and tubulins. Single and multiple amino acid replacements of Cct6p were constructed by oligonucleotide-directed mutagenesis, including changes of charged to alanine residues and uncharged to charged residues. The replacements were targeted, in part, to residues corresponding to functionally critical regions identified in the published crystal structure of the Escherichia coli chaperonin, GroEL. Here, we report the critical hydrophobic residues and clusters of hydrophilic residues in regions corresponding to those from the apical domain of GroEL implicated in peptide binding and peptide release, and certain residues in the putative equatorial domain implicated in subunit-to-subunit interaction. In contrast to their homologous counterparts in Cct2p and Cct1p, the highly conserved putative ATP binding motifs of Cct6p were relatively amenable to mutations. Our data suggest that the entire Cct6p molecule might be essential for assembly of Cct complex and might participate in binding substrates. However, there appeared to exist a functional hierarchy in ATP binding/hydrolysis among Cct subunits, as suggested by the high tolerance of Cct6p to mutations within the putative ATP binding pocket.

The effect of structural constraints in histidine ligation preference was investigated through studies of prepared two horse cytochrome c mutants, H26Q and H33N. Complementary equilibrium and stopped-flow kinetic experiment results revealed that His33 was the dominant heme iron ligand in GuHCl-unfolded cytochrome c. In the absence of His33, some other intramolecular ligand can still bind to the heme iron and can compete with His26.

The properties of an amino acid sequence that is needed for the trimethylation of lysine 72 in cytochrome c derived from Saccharomyces cerevisiae were evaluated by studying 21 altered iso-1-cytochromes c bearing single replacements in the region encompassing residues 67 through 77. The findings indicated that tyrosine 74 is critical for the trimethylation of lysine 72 while substitutions at other positions did not yield significant diminutions of the activity. The results further indicated that a properly situated aromatic amino acid, determined by the overall conformation fo apocytochrome c in the vicinity of lysine 72, appears to be important for trimethylation.

The signals required for forming 3[prime]-ends of mRNAs from the yeast Saccharomyces cerevisiae differ from the corresponding signals of higher eukaryotes. Yeast signals consist of three elements: (1) the efficiency element, which enhances the efficiency of downstream positioning elements; (2) the positioning element, which positions the poly(A) site; and (3) the actual poly(A) site. These three elements are not only necessary, but also sufficient for mRNA 3[prime]-end formation in yeast.

The role of helix pairing interactions is examined in relation to the folding process of cytochrome c proteins. This was done by using horse cytochrome c as a model. To gain added insight into the energetic and structural basis of these helix pairing interactions, the residue Leu94 was substituted with Ile, Val or Ala in a series of cytochrome c variants. Results show that Ile and Val substitutions led to slight changes in stability. In contrast, the Ala substitution led to a sharp decline in unfolding free energy.

Cytochrome c from the yeast Saccharomyces cerevisiae were modified to study the changes in electron transfer reaction rates caused by these modifications. Several cytochrome c isoforms were created that carried substitutions at key amino acids. The isoforms exhibited varying electron transfer rates that reflect their different free energy of reactions. This study illustrates one method of predicting changes in enzyme reaction rates in vivo and in vitro.

A study was conducted to test the hypothesis that B lymphocytes specific for self antigens (Ags) are also involved in the immune response to foreign Ags. Arg-specific monoclonal antibodies (mAbs) specific for pigeon cytochrome c (CYT) were compared with the mAbs for mouse CYT. B cells specific for mouse CYT responded only to pigeon CYT. The response became more specific as the response progressed, suggesting that the B cells in the primary response were also recruited to the secondary response.

A study was conducted to determine the molecular basis of the global suppressor activity for the N521 mutation. Isogenic strains were prepared using oligonucleotide-directed mutagenesis for the 20 amino acids at position 52 of iso-1-cytochrome c in the yeast Saccharomyces cerevisae. Results showed that the thermodynamic stabilization at position 52 is linked to the side chain hydrophobicity at the site.

A study was conducted to analyze the stability variants of cytochrome c in vitro. A C102A mutation was prepared in iso-1-cytochrome c. Oligonucleotides with mixed codons at the target site were used to produce the functional variants. The replacements enabled the study of the relationship between thermostability and proteolytic degradation of cytochromes c in the intermitochondrial space.

A study of the role of Arg38, Asn52 and Phe82 and the synergistic effects on the protein structure reveled that the functional and structural properties were affected in different ways when multiple mutations were introduced within a single protein, yeast iso-1 cytochrome c. For some cases, the effects were strictly additive while for the others they were synergistic. The stability effects of the three mutation sites were independent and cumulative in multiple-mutant proteins which reflects their independent nature. The difficulty of predicting synergistic effects of multiple mutations within a protein are underlined.