Predicting coiled coils from protein sequences.

The probability that a residue in a protein is part of a coiled-coil structure was assessed by comparison of its flanking sequences with sequences of known coiled-coil proteins. This method was used to delineate coiled-coil domains in otherwise globular proteins, such as the leucine zipper domains in transcriptional regulators, and to predict regions of discontinuity within coiled-coil structures, such as the hinge region in myosin. More than 200 proteins that probably have coiled-coil domains were identified in GenBank, including alpha- and beta-tubulins, flagellins, G protein beta subunits, some bacterial transfer RNA synthetases, and members of the heat shock protein (Hsp70) family.

Roles of the highly conserved aspartate and lysine residues in the response regulator of bacterial chemotaxis.

The chemotactic responses of bacteria such as Escherichia coli and Salmonella typhimurium are mediated by phosphorylation of the CheY protein. Phospho-CheY interacts with the flagellar motor switch to cause tumbly behavior. CheY belongs to a large family of phosphorylated response regulators that function in bacteria to control motility and regulate gene expression.

Urinary cyclic AMP excretion is increased in Macaca fascicularis and Macaca mulatta compared to humans.

Changes in parathyroid hormone and its second messenger cyclic AMP have been implicated in the pathogenesis of osteoporosis. Nonhuman primate models have been useful in the study of osteoporosis, but the physiology of mineral metabolism in certain species is different than in humans. We investigated parameters of mineral metabolism in 15 normal adult female cynomolgus and 14 normal adult female rhesus monkeys.

A comparison of the efficacy and safety of ramipril and digoxin added to maintenance diuretic treatment in patients with chronic heart failure.

The efficacy and safety of ramipril were compared with that of digoxin in a prospective, randomized, double-blind, crossover study of 35 patients with congestive heart failure (CHF), New York Heart Association (NYHA) grades II to IV, stabilized on diuretic maintenance therapy. Major assessments were conducted at baseline and at the end of each 10-week treatment period: primary efficacy variables were total exercise duration (modified Bruce, treadmill), NYHA grade, and clinical signs and symptoms (by visual analogue score) of heart failure. Twenty-seven patients completed the study.

Evidence for an S-farnesylcysteine methyl ester at the carboxyl terminus of the Saccharomyces cerevisiae RAS2 protein.

The protein products of yeast and mammalian ras genes are posttranslationally modified to give mature forms that are localized to the inner surface of the plasma membrane. We have previously demonstrated that the mature form of the Saccharomyces cerevisiae RAS2 gene product is methyl esterified at a modified C-terminal cysteine residue. Here we provide evidence that this residue is an S-farnesylcysteine alpha-carboxyl methyl ester.

Novel mutations that alter the regulation of sporulation in Bacillus subtilis. Evidence that phosphorylation of regulatory protein SpoOA controls the initiation of sporulation.

Sporulation in Bacillus subtilis is a complex developmental process that occurs in response to nutrient deprivation. To identify components of the mechanism that allows cells to monitor their nutritional status and to understand how this sensory information is transduced into a signal to activate specific sporulation genes, we have isolated mutants that are able to sporulate efficiently under nutritional conditions that strongly inhibit sporulation in wild-type bacteria, a phenotype we refer to as Coi (control of initiation). Four coi mutations were found to be within the coding sequence of spoOA, a gene in which null mutations prevent the initiation of sporulation and a gene whose product shares a domain of homology with phosphorylation-activated proteins that play signal transduction roles in bacteria.

Weight training decreases vertebral bone density in premenopausal women: a prospective study.

The effect of exercise on bone mass is unclear. To determine the skeletal effect of weight-bearing exercise in premenopausal women, we prospectively evaluated the effects of a weight-training program on lumbar spine bone mass in 10 women (mean +/- SEM, 36.2 +/- 1.

Cis-acting sequences from a human surfactant protein gene confer pulmonary-specific gene expression in transgenic mice.

Pulmonary surfactant is produced in late gestation by developing type II epithelial cells lining the alveolar epithelium of the lung. Lack of surfactant at birth is associated with respiratory distress syndrome in premature infants. Surfactant protein C (SP-C) is a highly hydrophobic peptide isolated from pulmonary tissue that enhances the biophysical activity of surfactant phospholipids.

Divalent metal ion binding to the CheY protein and its significance to phosphotransfer in bacterial chemotaxis.

Signal transduction in bacterial chemotaxis involves transfer of a phosphoryl group between the cytoplasmic proteins CheA and CheY. In addition to the established metal ion requirement for autophosphorylation of CheA, divalent magnesium ions are necessary for the transfer of phosphate from CheA to CheY. The work described here demonstrates via fluorescence studies that CheY contains a magnesium ion binding site.

Signal transduction in bacteria.

Cells display a remarkable ability to respond to small fluctuations in their surroundings. In simple microbial systems, information from sensory receptors feeds into a circuitry of regulatory proteins that transfer high energy phosphoryl groups from histidine to aspartate side chains. This phosphotransfer network couples environmental signals to an array of response elements that control cell motility and regulate gene expression.

Retinal vasculopathy associated with systemic light chain deposition disease.

A 35-year-old black male with the abrupt onset of blurred vision and decreased visual acuity was found to have severe retinal vasculopathy associated with systemic light chain deposition disease. While ocular manifestations are frequently reported in other plasma cell dyscrasias; ophthalmologic symptoms have not been previously reported in systemic light chain deposition disease alone. A possible mechanism of light chain deposition and damage to the retinal vasculature is proposed.

Protein phosphorylation and regulation of adaptive responses in bacteria.

Bacteria continuously adapt to changes in their environment. Responses are largely controlled by signal transduction systems that contain two central enzymatic components, a protein kinase that uses adenosine triphosphate to phosphorylate itself at a histidine residue and a response regulator that accepts phosphoryl groups from the kinase. This conserved phosphotransfer chemistry is found in a wide range of bacterial species and operates in diverse systems to provide different regulatory outputs.

Phosphorylation and dephosphorylation of a bacterial transcriptional activator by a transmembrane receptor.

Signal transduction in the bacterial Omp, Che, and Ntr systems involves the phosphorylation and dephosphorylation of response regulators (OmpR, CheY and CheB, NRI) that share a homologous domain. We show that in the Omp system, the transmembrane sensor EnvZ, catalyzes both the phosphorylation of OmpR and the dephosphorylation of OmpR-P. The phosphorylation reaction proceeds by a mechanism shared with the Ntr and Che kinases, NRII, and CheA.

Phosphorylation of an N-terminal regulatory domain activates the CheB methylesterase in bacterial chemotaxis.

Two types of reversible protein modification reactions have been identified in bacterial chemotaxis, methylation of membrane receptor-transducer proteins at glutamate side chains and phosphorylation of cytoplasmic signal transduction proteins at histidine and aspartate side chains. CheB is a bifunctional enzyme that is involved in both these modification processes. Its C-terminal domain is a methylesterase that catalyzes the hydrolysis of gamma-carboxyl glutamyl methyl esters in the cytoplasmic domain of chemoreceptor proteins.

Calcium-dependent release of N-terminal fragments and intact immunoreactive parathyroid hormone by human pathological parathyroid tissue in vitro.

The regulation of PTH secretion by extracellular calcium was studied in parathyroid tissue obtained from patients with hyperparathyroidism (adenoma or hyperplasia) using both an amino (N)-terminal RIA as well as an immunoradiometric assay (intact assay) specific for the intact hormone. The parathyroid glands separated into three major groups when examined in terms of absolute amounts of PTH secreted, degree of suppressibility, and set-point for calcium (the concentration of calcium causing half-maximal inhibition of PTH release). In cell preparations from group A (two different adenomas, two hyperplastic glands from a patient with renal failure, and a hyperplastic gland from a patient with hypophosphatemic rickets), both assays showed comparable PTH release (agreeing within 2-fold), similar degrees of suppressibility and similar, if not identical, set-points.

Evidence for a complex regulatory array in the first intron of the human adenosine deaminase gene.

Adenosine deaminase (ADA) is expressed ubiquitously by diverse mammalian cells and tissues but at levels that vary according to tissue and species. In humans, the thymus exhibits levels of the enzyme up to 100-fold higher than most other tissues. Using transgenic mice, we identified human ADA gene regulatory domains.

RAS2 protein of Saccharomyces cerevisiae is methyl-esterified at its carboxyl terminus.

Yeast and mammalian RAS gene products are GTP-binding proteins that are posttranslationally localized to the inner surface of the plasma membrane. This localization is accomplished by the addition of a lipid moiety to a conserved cysteine residue close to the carboxyl terminus. In a previous report we showed that the mammalia Ha-ras protein is also modified posttranslationally by methyl esterification.

Targeted ablation of alpha-crystallin-synthesizing cells produces lens-deficient eyes in transgenic mice.

Genetic ablation techniques were used to study the role of the lens in mammalian eye development. Ablation was accomplished by microinjecting murine eggs with chimeric DNA constructs in which the alpha A-crystallin gene regulatory sequence (-366 to +46) was fused to the highly cytotoxic diphtheria toxin gene coding sequence. For genetic ablation to be successful the promoter regulating expression should be specific and completely silent in cells necessary for normal mouse development.

Three-dimensional structure of CheY, the response regulator of bacterial chemotaxis.

Homologies among bacterial signal transduction proteins suggest that a common mechanism mediates processes such as chemotaxis, osmoregulation, sporulation, virulence, and responses to nitrogen, phosphorous and oxygen deprivation. A common kinase-mediated phosphotransfer reaction has recently been identified in chemotaxis, nitrogen regulation, and osmoregulation. In chemotaxis, the CheA kinase passes a phosphoryl group to the cytoplasmic protein CheY, which functions as a phosphorylation-activated switch that interacts with flagellar components to regulate motility.

Effects of estrogen in vivo and in vitro on spontaneous interleukin-1 release by monocytes from postmenopausal women.

Estrogen (E) inhibits bone resorption, but the mechanism of this effect is unknown. Interleukin-1 (IL-1) stimulates bone resorption in vitro and may be produced in bone by mononuclear phagocytes. Recently, the spontaneous release of IL-1 from peripheral monocytes was found to reflect bone formation in a subset of patients with idiopathic osteoporosis.

Calcitonin inhibits accumulation of cyclic AMP in stimulated peritoneal macrophages from normal rats but not from osteopetrotic (incisors-absent) littermates.

Macrophages and osteoclasts derive from related cell lines. In osteopetrotic mutants the function of osteoclasts is greatly reduced compared to that in normal animals or children and macrophage function is variably affected depending upon the mutation. To further explore macrophage function in osteopetrosis we examined the regulation of cyclic AMP production in macrophages from mutants and normal littermates of the osteopetrotic stock incisors-absent (ia) in the rat.