Herpes simplex virus 1 protein kinase is encoded by open reading frame US3 which is not essential for virus growth in cell culture.

Earlier reports have described a novel protein kinase in cells infected with herpes simplex or pseudorabies viruses. These novel enzymes were characterized by their acceptance of protamine as a substrate and by their differential chromatographic behavior in anion-exchange chromatography. We report that this activity was not present in extracts of uninfected cells or of cells infected with a mutant constructed so as to contain a deletion in the US3 open reading frame mapping in the small component of herpes simplex virus 1 DNA. The activity was present in extracts of cells infected with wild-type virus and with a recombinant in which the US3 open reading frame had been rescued. Our results are consistent with the observation reported earlier that the coding sequences predict an amino acid motif common to protein kinases and lead to the conclusion that the US3 open reading frame encodes a virus-specific protein kinase that is not required for virus growth in cells in culture.     

Site-directed mutagenesis of the cytoplasmic domains of the human beta 2-adrenergic receptor. Localization of regions involved in G protein-receptor coupling

Numerous plasma membrane-bound receptors are coupled to various effectors via a family of guanine nucleotide regulatory proteins (G proteins). Amino acid sequences of these receptors, deduced from cDNA and genomic clones, indicate the presence of seven transmembrane-spanning domains. Alignment of the available amino acid sequences of these G protein-linked receptors reveals striking homologies in regions predicted to lie near the cytoplasmic surface of the cell membrane. As these areas are likely those which interact with G proteins, we reasoned that systematic introduction of non-native sequence into these highly conserved regions of the human beta 2-adrenergic receptor would allow resolution of loci participating directly in receptor-G protein coupling. Based on this strategy, we constructed 19 mutant receptor species comprising substitutions and deletions of native sequence in the putative cytoplasmic domains of human beta 2-adrenergic receptor. By monitoring ligand binding characteristics and receptor-mediated stimulation of adenylyl cyclase, we have determined that the C-terminal portion of the third cytoplasmic loop and the N-terminal segment of the cytoplasmic tail appear to be critical for productive receptor-coupling to G proteins. In addition, we have implicated two other areas of the receptor that possibly play supportive roles in maintaining proper orientation of the G protein binding site. These comprise the second cytoplasmic loop and a conserved cysteine residue in the cytoplasmic tail.     

Duplicated region of the mouse genome containing a cytoplasmic gamma-actin processed pseudogene associated with long interspersed repetitive elements

The structures of two cloned recombinants of bacteriophage lambda and mouse genomic DNA (lambda mA14 and lambda mA36) were compared by electron microscopic analysis of various heteroduplex DNAs, restriction endonuclease mapping and nucleotide sequence determination. Each clone was shown to be derived from a distinct region of the mouse genome, but the two exhibited structural similarity over a region of at least 11,000 bases which included a cytoskeletal gamma-actin processed pseudogene of approximately 1800 bases. It is concluded that the two genomic regions were derived from a common ancestral region by duplication or amplification. The homologous regions of the two clones contained members of the long interspersed repetitive L1Md (long interspersed repeated sequence 1 of Mus domesticus) family lying in opposite orientation to one another, so that single-stranded DNA from the clones could form intra-molecular heteroduplexes. The complete nucleotide sequences of three L1Md members in lambda mA14 were determined. The longest of these (L1Md-14LH) had inserted into the gamma-actin processed pseudogene and, although it contained internal deletions, appeared to possess intact 5' and 3' ends. A second L1Md member (L1Md-14RH1) also appeared to have an intact 5' end but had lost most of its 3' portion, and a third member (L1Md-14RH2) was an internal fragment. The repeated sequence at the 5' ends of L1Md-14LH and L1Md-14RH1 showed these to be members of the L1Md-A family.     

Partial purification and characterization of a new phosphoprotein kinase from cells infected with pseudorabies virus

Cytoplasmic fractions from normal baby hamster kidney fibroblasts and from fibroblasts infected with pseudorabies virus were fractionated by DEAE-cellulose chromatography and fractions assayed for protein kinase activity. In preparations from uninfected and infected cells protein kinase activities identified as casein kinase I and II, the two isoforms of the cyclic-AMP-dependent protein kinase, protein kinase C, and a presumed proteolytic fragment of protein kinase C were present in comparable amounts. However in infected cells a new protein kinase activity was detected, appearing about 4 h after infection and increasing during the following 6 h at least. This new protein kinase was purified 100-fold by high-performance gel-permeation and ion-exchange chromatography, and characterized. It has an apparent relative molecular mass of 68 000 on the basis of gel-permeation chromatography, and a sedimentation coefficient of 4.3 S. It catalysed the phosphorylation of serine residues of basic proteins in vitro, with protamine a better substrate than mixed histones; and used ATP (apparent Km = 60 microM), but not GTP, as phosphoryl donor. Molecules that can serve as effectors for other protein kinases (cyclic AMP, cyclic GMP, Ca2+ + calmodulin, Ca2+ + phospholipid, double-stranded RNA, and heparin) did not significantly alter the activity of this enzyme. A distinguishing characteristic of the protein kinase was a high KCl concentration optimum with the persistence of activity up to 800 mM KCl, at least.     

Maternal nutrition in pregnancy. Part I: a review.

Maternal undernutrition may result in a greater deprivation of the fetus than has previously been believed. The infant not only may be "light for dates" but also has an increased risk of perinatal disability or death secondary to gross neurologic and developmental abnormalities. This article reviews current knowledge of the energy, protein, iron, vitamin, sodium and calcium requirements in pregnancy, with special reference to the management of the underweight and overweight pregnant women.     

Increased phosphorylation of ribosomal protein S6 in hamster fibroblasts transformed by polyoma virus and simian virus 40

The extent of phosphorylation of ribosomal protein S6 was compared in normal hamster fibroblasts and in fibroblasts transformed by polyoma virus or simian virus 40. In both strains of transformed cells the protein was more highly phosphorylated than in the normal cells.