Addition of copper to yeast cells leads to the induction of a low molecular weight, cysteine-rich protein that binds copper. This protein, termed copper chelatin or thionein, is related to the metallothionein family of proteins that are induced in response to cadmium and zinc in vertebrate cells. We have determined the structure of the yeast copperbinding protein by DNA sequence analysis of the gene. Although the 6573-dalton yeast protein is substantially divergent from vertebrate metallothioneins, the arrangement of 12 cysteine residues, which is a hallmark of metal-binding proteins, is partially conserved. We analyzed the regulatory DNA sequence of the gene by fusing it with the Escherichia coli galactokinase gene and assaying the levels of enzyme activity in yeast in response to copper. The transcriptional activation has a specific requirement for copper. Zinc, cadmium, and gold were unable to regulate the galactokinase activity. The yeast copper metallothionein regulatory sequences represent a previously unreported class of yeast promoter that is regulated by copper.Methallothioneins (MTs) are thought to play a central role in the protection against heavy metal toxicity and zinc homeostasis (see ref. 1 for a review). Recent interest in the molecular biology of MT stems from the observation that the genes for mammalian MT are regulated by heavy metals as well as by glucocorticoids (2-3). MT genes from several different mammalian systems such as mouse, Chinese hamster, monkey, and human have been cloned and sequenced (4-7).Although the precise mechanism of MT gene activation by heavy metals has not been elucidated, a DNA sequence upstream of the 5' end of the mouse MT I gene, which is responsible for cadmium inducibility, has been identified (8, 9 Recently we cloned a copper-inducible gene from a CUpir strain and showed that it protects a cupP strain from copper toxicity (14). The restriction map of this clone indicated that it is identical to the CUP] gene previously cloned and characterized by Fogel and Welch (12). To provide a basis for understanding the function and regulation of this gene, we determined the primary structure of the coding region and flanking sequences. We show that it encodes a protein with both similarities to and differences from the MTs of higher eukaryotes. We refer to this protein as copper metallothionein (Cu-MT) because it belongs to the metallothionein family of proteins as defined in ref. Fig. 4). Growth 3332The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Penciclovir (PCV), an antiherpesvirus agent in the same class as acyclovir (ACV), is phosphorylated in herpes simplex virus (HSV)-infected cells by the viral thymidine kinase (TK).Resistance to ACV has been mapped to mutations within either the TK or the DNA polymerase gene. An identical activation pathway, the similarity in mode of action, and the invariant cross-resistance of TK-negative mutants argue that the mechanisms of resistance to PCV and ACV are likely to be analogous. A total of 48 HSV type 1 (HSV-1) and HSV-2 isolates were selected after passage in the presence of increasing concentrations of PCV or ACV in MRC-5 cells. Phenotypic analysis suggested these isolates were deficient in TK activity. Moreover, sequencing of the TK genes from ACV-selected mutants identified two homopolymeric G-C nucleotide stretches as putative hot spots, thereby confirming previous reports examining Acv r clinical isolates. Surprisingly, mutations identified in PCV-selected mutants were generally not in these regions but distributed throughout the TK gene and at similar frequencies of occurrence within A-T or G-C nucleotides, regardless of virus type. Furthermore, HSV-1 isolates selected in the presence of ACV commonly included frameshift mutations, while PCV-selected HSV-1 mutants contained mostly nonconservative amino acid changes. Data from this panel of laboratory isolates show that Pcv r mutants share cross-resistance and only limited sequence similarity with HSV mutants identified following ACV selection. Subtle differences between PCV and ACV in the interaction with viral TK or polymerase may account for the different spectra of genotypes observed for the two sets of mutants.The introduction of penciclovir [PCV;9-(4-hydroxy-3-hydroxymethylbut-1-yl)guanine] and its prodrug, famciclovir, (FCV), resulted in the use of antivirals alternative to acyclovir (ACV) for treatment of herpes simplex virus (HSV) infections. Biochemical studies have indicated that PCV, like ACV, is phosphorylated by the viral thymidine kinase (TK) to a monophosphate and subsequently converted by cellular enzymes to a triphosphate, which inhibits the HSV DNA polymerase (Pol) (44). Although PCV and ACV have identical activation pathways and similar modes of action (14, 44), and the frequencies with which resistance in HSV arises to PCV and ACV in cell culture are identical (36), the affinities and therefore the fine molecular interactions of PCV, ACV, and their triphosphates with TK and Pol differ (14). The last point raises the possibility that drug-resistant mutants selected by these antiviral agents may differ.Resistance to acyclovir typically arises by a single mutation in either the TK or Pol gene (11,23,29). The viral TK, unlike DNA polymerase, is not essential for virus replication in cell culture (13), although in vivo analyses implicate it in HSV virulence, pathogenicity, and reactivation from latency (9,15,20,41). Mutations in HSV TK are the most common causes of clinical resistance to ACV (7,34), and the majority of mutants completely lac...

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There are a variety of databases relevant to nursing practice and nursing research. Access to databases such as Medline and Eric has largely been through libraries or other institutions, but with the advent of personal computers it is now possible for anyone to access a database. This article discusses the advantages of personally conducting computerized literature searches. The hardware and software necessary to conduct a computer search are also reviewed.