Reduction in postprandial energy expenditure during pregnancy.

Energy expenditure was measured during pregnancy in seven primigravid women at 12-15, 25-28, and 34-36 weeks and after the cessation of lactation. On each occasion the resting metabolic rate and the increase in metabolic rate after ingestion of a liquid test meal were measured by indirect calorimetry. In absolute terms the resting metabolic rate increased steadily during pregnancy but when expressed per unit of body weight no change was found. The energetic response to a mixed constituent meal was significantly reduced by 28% in the middle trimester of pregnancy. These findings suggest a possible maternal adaptation to increase energetic efficiency at a time when the energy demands of the fetus are high.     

The Drosophila tissue polarity gene starry night encodes a member of the protocadherin family.

The tissue polarity genes control the polarity of hairs, bristles and ommatidia in the adult epidermis of Drosophila. We report here the identification of a new tissue polarity gene named starry night (stan). Mutations in this essential gene alter the polarity of cuticular structures in all regions of the adult body. The detailed polarity phenotype of stan on the wing suggested that it is most likely a component of the frizzled (fz) pathway. Consistent with this hypothesis, stan appears to be downstream of and required for fz function. We molecularly cloned stan and found that it encodes a huge protocadherin containing nine cadherin motifs, four EGF-like motifs, two laminin G motifs, and seven transmembrane domains. This suggests that Stan functions in signal reception, perhaps together with Fz.     

Substrate analogue induced changes of the CO-stretching mode in the cytochrome P450cam-carbon monoxide complex.

The CO-stretching mode of the carbon monoxide ligand in reduced cytochrome P450cam, in the absence or presence of camphor and in the presence of nine different camphor analogues, was measured at room temperature using Fourier transform infrared spectroscopy. Substrate-free cytochrome P450cam--CO reveals a broad, slightly structured band resulting from an overlap of several stretching mode signals. The multitude of the signals indicates that cytochrome P450 exists in a dynamic equilibrium of several conformational substates. Binding of camphor or camphor analogues strongly influences this equilibrium. For substrate analogues which are not able to form a hydrogen bond to the hydroxyl group of tyrosine 96, the CO-stretching band is rather broad and asymmetric. In contrast, substrate analogues with one quinone group which form a hydrogen bond to the Tyr96 OH induce a shift and a sharpening of the CO-stretching mode band. For substrate analogues with two hetero groups, the infrared spectrum is slightly asymmetric or a minor band appears. Sterical hindrance, substrate mobility, and protein flexibility finally determine the position and width of the CO-stretching mode signals.     

Free fatty acids activate a vigorous Ca(2+):2H(+) antiport activity in yeast mitochondria.

The accumulation and retention of Ca(2+) by yeast mitochondria (Saccharomyces cerevisiae) mediated by ionophore ETH 129 occurs with a variable efficiency in different preparations. Ineffective Ca(2+) transport and a depressed membrane potential occur in parallel, are exacerbated in parallel by exogenous free fatty acids, and are corrected in parallel by the addition of bovine serum albumin. Bovine serum albumin is not required to develop a high membrane potential when either Ca(2+) or ETH 129 are absent, and when both are present membrane potential is restored by the addition of EGTA in a concentration-dependent manner. Respiration and swelling data indicate that the permeability transition pore does not open in yeast mitochondria that are treated with Ca(2+) and ETH 129, whereas fatty acid concentration studies and the inaction of carboxyatractyloside indicate that fatty acid-derived uncoupling does not underlie the other observations. It is concluded that yeast mitochondria contain a previously unrecognized Ca(2+):2H(+) antiporter that is highly active in the presence of free fatty acids and leads to a futile cycle of Ca(2+) accumulation and release when exogenous Ca(2+) and ETH 129 are available. It is also shown that isolated yeast mitochondria degrade their phospholipids at a relatively rapid rate. The activity responsible is also previously unrecognized. It is Ca(2+)-independent, little affected by the presence or absence of a respiratory substrate, and leads to the hydrolysis of ester linkages at both the sn-1 and sn-2 positions of the glycerophospholipids. The products of this activity, through their actions on the antiporter, explain the variable behavior of yeast mitochondria treated with Ca(2+) plus ETH 129.     

Structural basis of human erythrocyte glucose transporter function in reconstituted system. Hydrogen exchange

Hydrogen exchange kinetic behavior of human erythrocyte glucose transporter protein in vesicles was studied in the absence and in the presence of D-glucose or a well known inhibitor, cytochalasin B. This is to detect a proposed channel of water penetrating into the protein through which the sugar molecule passes and to monitor any conformational changes induced by the substrate or inhibitor. Analyses of the kinetic data revealed several classes of hydrogens which exchange with readily distinguishable rates. Of 660 hydrogens detected per transporter, approximately 30% exchanged with rates generally characterized as those of free amide hydrogens indicating they are interfaced to solvent water. Since the transporter is known to be embedded deep in the hydrophobic area of the membrane with minimum exposure to the outside of the membrane lipid bilayer, a significant portion of these free amide hydrogens must be at the purported channel rather than outside of the membrane. D-Glucose and cytochalasin B affected the exchange kinetics of these presumably channel-associated free amide hydrogens rather differently. D-Glucose reduced the apparent rate constants, but not the total number. Cytochalasin B on the other hand reduced the total number to one-half without significantly changing the apparent rate constants. The remaining 70% of the labeled hydrogens exchanged with much slower rates which vary 10-10,000-fold, indicating that they are internally structured peptide amide and side chain hydrogens. Both D-glucose and cytochalasin B further reduced the rates of these hydrogens, indicating a global stabilization of the protein structure.     

Lipopeptides of the N-terminus of Escherichia coli lipoprotein: synthesis, mitogenicity and properties in monolayer experiments

The N-terminal part of the lipoprotein from the outer membrane of Escherichia coli, tripalmitoyl-S-glyceryl-L-Cys-Ser and analogs with longer sequences, are polyclonal activators for B-lymphocytes. Triple-chain lipopeptides also constitute efficient low-molecular-weight carrier/adjuvant systems, which can be linked to antigens to yield immunogens for antibody production without further additives. This is the first report of monolayer experiments with chemically well defined, synthetic lipopeptide mitogens with the composition of the N-terminus of an important bacterial membrane protein. Various derivatives of the lipoprotein N-terminus were synthesized. These lipopeptides differed in the length of the peptide moiety, the number of fatty acid residues, and protective groups. In order to obtain the surface areas for the lipopeptides in isotherms and hysteresis isotherms, monolayer experiments with a computer-controlled film balance were performed. To get some information about the interaction of these compounds with typical membrane lipids mixed monolayers were formed from triple-chain lipopeptides with dipalmitoylphosphatidylcholine and cholesterol. A comparison of the mitogenic response of the compounds was made in an in vitro system with B-lymphocytes from Balb/c mice.     

Electron Microscopic Studies of Spruce Needles in Connection with the Occurrence of Novel Forest Decline

Needles of four spruce trees showing different degrees of novel kinds of forest decline were investigated by electron microscopy. Green needles appearing at least superficially still intact were selected for the present investigation. Most of the mesophyll appeared to be undamaged. However, groups of atypical mesophyll cells were found close to the endodermis or the hypodermis. The chloroplasts of the apparently damaged cells were particularly affected. Changes in the matrix of the chloroplasts, i.e,. increased affinity to osmium, occurrence of extensive nests of plastoglobuli, as well as damage to the membranes, i.e. lesions in the envelope and abnormal thylakoid membranes, were observed. Signs of decomposition of other cellular structures including mitochondria were also detectable. There appeared to be a close correlation between the degree of damage at the whole tree level and the degree of damage occurring at the cellular level. It is concluded that particularly the lipids and the proteinsof, the membranes are affected by anthropogenic air pollutants and natural stressors. The altered membrane structure may for instance cause abnormal osmotic conditions for the cellular compartments and may impair transport processes and thus lead to lossof function not only of the cells but also of the whole needle.     

Expression and sequence analysis of a cDNA encoding the orotidine-5'-monophosphate decarboxylase domain from Ehrlich ascites uridylate synthase

Orotidine-5'-monophosphate decarboxylase (OD-Case) catalyzes the conversion of orotidine 5'-monophosphate to UMP. In mammals, ODCase is present as part of a bifunctional protein which also contains orotate phosphoribosyltransferase; the preceding enzyme in the de novo UMP biosynthetic pathway. We have isolated a plasmid (pMEJ) which contains a cDNA for the ODCase domain of UMP synthase. Insertion of this sequence into an Escherichia coli expression vector (pUC12) has allowed for the expression of ODCase and not orotate phosphoribosyltransferase in E. coli. The molecular weight of the expressed protein is 26,000-27,300 from immunoblot analysis which corresponds closely to the molecular weight of the ODCase domain (28,500) isolated by tryptic digestion of UMP synthase. We have sequenced the cDNA insert of pMEJ and deduced the amino acid sequence. The molecular weight of the ODCase domain calculated from the amino acid sequence in 28,654. Comparison of the deduced amino acid sequence from pMEJ with that for yeast ODCase (a monofunctional protein) demonstrated that 52% of the amino acids were identical when the two sequences are compared. Furthermore, several stretches of the amino acid sequence have 80% or greater absolute homology.