A critique of the utility of the prediction of protein secondary structure

The Chou-Fasman predictive algorithm for determining the secondary structure of proteins from the primary sequence is reviewed. Many examples of its use are presented which illustrate its wide applicability, such as predicting (a) regions with the potential for conformational change, (b) sequences which are capable of assuming several conformations in different environments, (c) effects of single amino acid mutations, (d) amino acid replacements in synthesis of peptides to bring about a change in conformation, (e) guide to the synthesis of polypeptides with definitive secondary structure,e.g. signal sequences, (f) conformational homologues from varying sequences and (g) the amino acid requirements for amphiphilicα-helical peptides.     

The histidine permease gene (HIP1) of Saccharomyces cerevisiae

The histidine-specific permease gene (HIP1) of Saccharomyces cerevisiae has been mapped, cloned, and sequenced. The HIP1 gene maps to the right arm of chromosome VII, approx. 11 cM distal to the ADE3 gene. The gene was isolated as an 8.6-kb BamHI-Sau3A fragment by complementation of the histidine-specific permease deficiency in recipient yeast cells. We sequenced a 2.4-kb subfragment of this BamHI-Sau3A fragment containing the HIP1 gene and identified a 1596-bp open reading frame (ORF). We confirmed the assignment of the 1596-bp ORF as the HIP1 coding sequence by sequencing a hip1 nonsense mutation. Analysis of the amino acid (aa) sequence of the HIP1 gene reveals several hydrophobic stretches, but shows no obvious N-terminal signal peptide. We have constructed a deletion of the HIP1 gene in vitro and replaced the wild-type copy of the gene with this deletion. The hip1 deletion mutant can grow when it is supplemented with 30 mM histidine, 50 times the amount required for the growth of HIP1 cells. Revertants of this deletion mutant able to grow on a normal level of histidine arise by mutation in unlinked genes. Both these observations suggest that there are additional, low-affinity pathways for histidine uptake.     

The trajectory of a stiff rod in a curved potential energy trough

The equilibrium trajectory of the axis of a rod subject to an externally imposed curved potential energy trough tends to conform to the shape of the curved trough, but also tends to be straight because of elastic resistance to bending. The actual path of the axis is a balance between the two extremes. We consider a potential energy trough centered along a circular arc of radiusR. For a rod of small length compared toR, we show that the axis at equilibrium forms an arc of a circle of radius greater thanR. The value of the radius of the axial path depends on the relative values of the Hooke’s Law bending constant for the rod and the depth and width of the trough. Motivation for the calculation is provided by nucleosomal DNA, which conforms to the surface of a roughtly cylindrical histone core at physiological ionic strength, but is observed to unwind into a partially extended conformation at very low ionic strength. We suggest that the rigidity to bending of short DNA segments becomes sufficiently great at low ionic strength to overcome attractive interactions with the histone surface. Alternately, of course, if during the cell cycle mutually attractive forces between DNA and histone core are weakened at constant ionic strength, the same type of unfolding would be expected to occur as the strength of the DNA-histone contacts drops below the level required to overcome elastic resistance to bending of the DNA rod.     

The influence of mineral salts on fecundity of the water flea (Daphnia magna) and the implications on toxicity testing of industrial wastewater

The effects of mineral salts constituting water hardness on fecundity ofDaphnia magna were assessed. Of the salts tested, increased concentrations of NaHCO₃ and MgSO₄ had no effect on fecundity, CaSO₄ significantly increased fecundity, and KCl significantly reduced fecundity. The number of offspring produced per daphnid was correlative to the CaSO₄ concentration at CaSO₄ concentrations between 91 and 2100 mg/ℓ. The effects of CaSO₄ on daphnid fecundity could influence the interpretive outcome of industrial wastewater toxicity tests using this species when the waste and dilution waters contain different concentrations of CaSO₄. It is recommended that when performing these tests, dilution water be sampled at the intake site of the industry's water source, thus assuring initial comparability of the waste and dilution waters. The CaSO₄ content of the water prior to and after industrial use should be determined to identify any alterations of CaSO₄ concentration during use. Identification of CaSO₄ concentration differences can aid in the interpretation of effects associated with the wastewater.     

Endogenous phosphorylation of retinal photoreceptor outer segment proteins by calcium phospholipid-dependent protein kinase

A calcium phospholipid-dependent protein kinase (C-kinase) activity was detected in the soluble fraction of rod outer segments (ROS) of the bovine retina. The enzyme required calcium, phosphatidylserine (PS) and diacylglycerol for maximal activity. In the presence of calcium and PS, C-kinase endogenously phosphorylated proteins with molecular weights of 95,000, 91,000, 31,000, 21,000, 19,000, 18,000, 16,000, 14,000 and 11,000. Addition of diolein in the reaction mixture further enhanced the endogenous phosphorylation of these proteins. Retinal was found to inhibit the phosphorylation of endogenous proteins by C-kinase in a concentration dependent manner. Half-maximal inhibition of enzyme activity was obtained at a retinal concentration of about 12μM. These results suggest that calcium, phospholipids and the C-kinase enzyme may play an important role in the functional regulation of rod photoreceptors and, with retinal, perhaps in the visual process as well.     

Inhibition of mast cell chymase by eglin c and antileukoprotease (HUSI-I). Indications for potential biological functions of these inhibitors

Recombinant eglin c (originally isolated from the medical leech) and antileukoprotease (HUSI-I from human seminal plasma) were examined for their ability to inhibit the mastcell protease chymase. Both inhibitors react rapidly with the enzyme: when about equimolar concentrations (in the range of 10(-8) M) of chymase and HUSI-I or eglin c were incubated the complex formation was apparently at equilibrium after 1 or 5 min respectively. When a constant amount of chymase (approximately 3 X 10(-8) M) was incubated with increasing concentrations of inhibitor a concentration of HUSI-I of 7 X 10(-7) M was necessary to cause 50% inhibition of the initial enzyme activity, whereas 8 X 10(-8) M eglin c was sufficient. The dissociation constant of the chymase-eglin c complex was calculated to be 4.4 X 10(-8) M. These results are discussed with respect to the possible in vivo function of antileukoprotease as an inhibitor of mast cell chymase.     

Saccharomyces cerevisiae nuclear fusion requires prior activation by alpha factor.

We have developed a protocol for efficient fusion of spheroplasts of the same mating type. Nuclear fusion in this whole-cell system is also efficient and closely parallels nuclear fusion in heterosexual mating of intact cells. In the spheroplast fusion system, nuclear fusion is dependent on both the KAR1 gene and prior exposure to alpha factor. The major products of nuclear fusion in the spheroplast fusion assay were true diploids that were homozygous at the mating-type locus. An additional 10% of the products were cells of ploidy greater than diploid. The dependence of nuclear fusion on alpha factor treatment could not be replaced by synchronization in G1 by mutations in CDC28 and CDC35 or by prior arrest in stationary phase. These data suggest that nuclear fusion is not a constitutive function of the nucleus, but rather is specifically induced by mating hormone.     

Histophotometric Evaluation of Glutamate Dehydrogenase Activity of the Rat Hippocampal Formation During Postnatal Development,with Special Reference to the Glutamate Transmitter Metabolism

Transmitter glutamate/aspartate synthesis is known to proceed along different metabolic pathways. In this light, the functional relevance of glutamate dehydrogenase in postnatally maturing glutamatergic/aspartatergic structures was studied by means of quantitative enzyme histochemistry. The basic requirements concerning the kinetics and calibration of the histochemical glutamate dehydrogenase reaction used were proved to be met in order to obtain valid quantitative data. The histochemically demonstrable activity of glutamate dehydrogenase (EC 1.4.1.3) in the hippocampal formation of the rat increased markedly during postnatal development. On day 30, the distribution pattern observed was similar to that in adult animals. While the enzyme activity rose within cell body layers from day 0 to day 30 by 240-285%, the increase in neuropil layers was found to be up to 830%. Maximum values were seen in the stratum lacunosum-moleculare of CA1 and CA3 and the stratum moleculare of the dentate fascia on day 30. Since the hippocampal neuropil is supposed to be copiously provided with glutamatergic (and aspartatergic?) structures which become functional in rats during the first weeks of postnatal life, the increase in enzyme activity is discussed to be primarily a consequence of maturing synaptic systems using glutamate and/or aspartate as transmitters.