Sequence of the alpha subunit of photoreceptor G protein: homologies between transducin, ras, and elongation factors

A bovine retinal complementary DNA clone encoding the alpha subunit of transducin (T alpha) was isolated with the use of synthetic oligodeoxynucleotides as probes, and the complete nucleotide sequence of the insert was determined. THe predicted protein sequence of 354 amino acids includes the known sequences of four tryptic peptides and sequences adjacent to the residues that undergo adenosine diphosphate ribosylation by cholera toxin and pertussis toxin. On the basis of homologies to other proteins, such as the elongation factors of protein synthesis and the ras oncogene proteins, regions are identified that are predicted to be acylated and involved in guanine nucleotide binding and hydrolysis. Amino acid sequence similarity between T alpha and ras is confined to these regions of the molecules.     

The DNA-binding properties of the major regulatory protein alpha 4 of herpes simplex viruses

The transition from the expression of alpha, the first set of five herpes simplex virus genes expressed after infection, to beta and gamma genes, expressed later in infection, requires the participation of infected cell protein 4 (alpha 4), the major viral regulatory protein. The alpha 4 protein is present in complexes formed by proteins extracted from infected cells and viral DNA fragments derived from promoter domains. This report shows that the alpha 4 protein forms specific complexes with DNA fragments derived from 5' transcribed noncoding domains of late (gamma 2) genes whose expression requires viral DNA synthesis as well as functional alpha 4 protein. Some of the DNA fragments to which alpha 4 binds do not contain homologs of the previously reported DNA binding site consensus sequence, suggesting that alpha 4 may recognize and interact with more than one type of DNA binding site. The alpha 4 proteins can bind to DNA directly. A posttranslationally modified form of the alpha 4 protein designated alpha 4c differs from the alpha 4a and alpha 4b forms with respect to its affinity for DNA fragments differing in the nucleotide sequences of the binding sites.     

A G protein gamma subunit shares homology with ras proteins

Guanine nucleotide binding proteins (G proteins) that transduce signals from cell surface receptors to effector molecules are made up of three subunits, alpha, beta, and gamma. A complementary DNA clone that encodes a 71-amino acid protein was isolated from bovine brain; this protein contains peptide sequences that were derived from the purified gamma subunit of Gi and Go. The primary sequence of this G protein gamma subunit (G gamma) has 55 percent homology to the gamma subunit of transducin (T gamma) and also has homology to functional domains of mammalian ras proteins. The probe for isolating the clone was generated with the use of the polymerase chain reaction (PCR). The extent of divergence between T gamma and G gamma, the isolation of homologous PCR-generated fragments, and the differences between the predicted amino acid sequence of G gamma and that derived from the gamma subunit of Gi and Go indicate that gamma subunits are encoded by a family of genes.     

The primary structure and heterogeneity of tau protein from mouse brain

Tau protein is a family of microtubule binding proteins, heterogeneous in molecular weight, that are induced during neurite outgrowth and are found prominently in neurofibrillary tangles in Alzheimer's disease. The predicted amino acid sequences of two forms of tau protein from mouse brain were determined from complementary DNA clones. These forms are identical in their amino-terminal sequences but differ in their carboxyl-terminal domains. Both proteins contain repeated sequences that may be tubulin binding sites. The sequence suggests that tau is an elongated molecule with no extensive alpha-helical or beta-sheet domains. These complementary DNAs should enable the study of various functional domains of tau and the study of tau expression in normal and pathological states.     

Methylation-sensitive sequence-specific DNA binding by the c-Myc basic region

The function of the c-Myc oncoprotein and its role in cell growth control is unclear. A basic region of c-Myc is structurally related to the basic motifs of helix-loop-helix (HLH) and leucine zipper proteins, which provide sequence-specific DNA binding function. The c-Myc basic region was tested for its ability to bind DNA by attaching it to the HLH dimerization interface of the E12 enhancer binding factor. Dimers of the chimeric protein, termed E6, specifically bound an E box element (GGCCACGTGACC) recognized by other HLH proteins in a manner dependent on the integrity of the c-Myc basic motif. Methylation of the core CpG in the E box recognition site specifically inhibited binding by E6, but not by two other HLH proteins. Expression of E6 (but not an E6 DNA binding mutant) suppressed the ability of c-myc to cooperate with H-ras in a rat embryo fibroblast transformation assay, suggesting that the DNA recognition specificity of E6 is related to that of c-Myc in vivo.     

Molecular cloning of complementary DNA for the alpha subunit of the G protein that stimulates adenylate cyclase

A complementary DNA clone encoding the alpha subunit of the adenylate cyclase stimulatory G protein (Gs) was isolated and identified. A bovine brain complementary DNA library was screened with an oligonucleotide probe derived from amino acid sequence common to known G proteins. The only clone that was obtained with this probe has a complementary DNA insert of approximately 1670 base pairs. An antibody to a peptide synthesized according to deduced amino acid sequence reacts specifically with the alpha subunit of Gs. In addition, RNA that hybridizes with probes made from the clone is detected in wild-type S49 cells; however, cyc- S49 cells, which are deficient in Gs alpha activity, are devoid of this messenger RNA.     

Homeo boxes in the study of development

The body plan of Drosophila is determined to a large extent by homeotic genes, which specify the identity and spatial arrangement of the body segments. Homeotic genes share a characteristic DNA segment, the homeo box, which encodes a defined domain of the homeotic proteins. The homeo domain seems to mediate the binding to specific DNA sequences, whereby the homeotic proteins exert a gene regulatory function. By isolating the normal Antennapedia gene, fusing its protein-coding sequences to an inducible promoter, and reintroducing this fusion gene into the germline of flies, it has been possible to transform head structures into thoracic structures and to alter the body plan in a predicted way. Sequence homologies suggest that similar genetic mechanisms may control development in higher organisms.     

A single receptor binds both insulin-like growth factor II and mannose-6-phosphate

Amino acid sequences deduced from rat complementary DNA clones encoding the insulin-like growth factor II (IGF-II) receptor closely resemble those of the bovine cation-independent mannose-6-phosphate receptor (Man-6-P receptorCI), suggesting they are identical structures. It is also shown that IGF-II receptors are adsorbed by immobilized pentamannosyl-6-phosphate and are specifically eluted with Man-6-P. Furthermore, Man-6-P specifically increases by about two times the apparent affinity of the purified rat placental receptor for 125I-labeled IGF-II. These results indicate that the type II IGF receptor contains cooperative, high-affinity binding sites for both IGF-II and Man-6-P-containing proteins.     

Convergence of Ets- and notch-related structural motifs in a heteromeric DNA binding complex

Analysis of the heteromeric DNA binding protein GABP has revealed the interaction of two distinct peptide sequence motifs normally associated with proteins located in different cellular compartments. The alpha subunit of GABP contains an 85-amino acid segment related to the Ets family of DNA binding proteins. The ETS domain of GABP alpha facilitates weak binding to DNA and, together with an adjacent segment of 37 amino acids, mediates stable interaction with GABP beta. The beta subunit of GABP contains four imperfect repeats of a sequence present in several transmembrane proteins including the product of the Notch gene of Drosophila melanogaster. These amino-terminal repeats of GABP beta mediate stable interaction with GABP alpha and, when complexed with GABP alpha, directly contact DNA. These observations provide evidence for a distinct biochemical role for the 33-amino acid repeats, and suggest that they may serve as a module for the generation of specific dimerization interfaces.     

Biosynthesis of the Torpedo californica acetylcholine receptor alpha subunit in yeast

Yeast cells were transformed with a plasmid containing complementary DNA encoding the alpha subunit of the Torpedo californica acetylcholine receptor. These cells synthesized a protein that had the expected molecular weight, antigenic specificity, and ligand-binding properties of the alpha subunit. The subunit was inserted into the yeast plasma membrane, demonstrating that yeast has the apparatus to express a membrane-bound receptor protein and to insert such a foreign protein into its plasma membrane. The alpha subunit constituted approximately 1 percent of the total yeast membrane. The alpha subunit constituted approximately 1 percent of the total yeast membrane proteins, and its density was about the same in the plasma membrane of yeast and in the receptor-rich electric organ of Electrophorus electricus. In view of the available technology for obtaining large quantities of yeast proteins, it may now be possible to obtain amplified amounts of interesting membrane-bound proteins for physical and biochemical studies.     

Genetic variation in the human insulin gene

Four recombinant lambda phages containing nucleotide sequences complementary to a cloned human preproinsulin DNA probe have been isolated from human DNA. Restriction analyses in conjunction with Southern hybridizations reveal two types of gene sequences. One isolate of each type was subjected to complete nucleotide sequence determination. The sequences contain the entire preproinsulin messenger RNA region, two intervening sequence. 260 nucleotides upstream from the messenger RNA capping site, and 35 nucleotides beyond the polyadenylate attachment site. Our results strongly suggest that these two gene types are allelic variants of a single insulin gene.