Structural Comparisons of Muscle and Nonmuscle Actins Give Insights Into the Evolution of Their Functional Differences

Actin is a highly conserved protein although many isoforms exist. In vertebrates and insects the different actin isoforms can be grouped by their amino acid sequence and tissue-specific gene expression into muscle and nonmuscle actins, suggesting that the different actins may have a functional significance. We ask here whether atomic models for G- and F-actins may help to explain this functional diversity. Using a molecular graphics program we have mapped the few amino acids that differ between isoactins. A small number of residues specific for muscle actins are buried in internal positions and some present a remarkable organization. Within the molecule, the replacements observed between muscle and nonmuscle actins are often accompanied by compensatory changes. The others are dispersed on the protein surface, except for a cluster located at the N-terminus which protrudes outward. Only a few of these residues specific for muscle actins are present in known ligand binding sites except the N-terminus, which has a sequence specific for each isoactin and is directly implicated in the binding to myosin. When we simulated the replacements of side chains of residues specific for muscle actins to those specific for nonmuscle actins, the N-terminus appears to be less compact and more flexible in nonmuscle actins. This would represent the first conformational grounds for proposing that muscle and nonmuscle actins may be functionally distinguishable. The rest of the molecule is very similar or identical in all the actins, except for a possible higher internal flexibility in muscle actins. We propose that muscle actin genes have evolved from genes of nonmuscle actins by substitutions leading to some conformational changes in the protruding N-terminus and the internal dynamics of the main body of the protein. Received: 15 March 1996 / Accepted: 14 July 1996     

VIP receptors in pig liver cells: binding characteristics and role in degradation

The physiological role of VIP in the liver is controversial. VIP receptors are present, but their function in the metabolic regulation is uncertain. The interaction of porcine VIP with isolated cells from pig liver was studied with respect to receptor-binding, degradation and glycogenolytic action. In this model, VIP and liver showed homology of animal species. 1. Receptor-binding was heterogenous with Kd values of 10(-9) mol/l and 4 X 10(-8) mol/l, and a total amount of binding sites of 7 X 10(-11) mol per 10(9) cells. The peptide specificity showed that porcine and chicken VIP were equally potent in inhibiting receptor-bound 125I-VIP; secretin was about 30 times less potent; glucagon and somatostatin were ineffective. 2. Receptor-bound 125I-VIP was degraded since about 70% was released as radioactivity not reacting with VIP-antiserum. 3. Glucose-release was not stimulated by VIP (10(-6) mol/l) whereas the rate was increased two-fold by glucagon (10(-6) mol/l). In conclusion, VIP receptors in pig liver cells are different from other tissues regarding peptide specificity. It is suggested that receptor-binding mediates degradation of VIP by pig liver rather than metabolic effects.     

Analysis of penicillin-binding sites with a micro method — The binding site of PBP 5 from Escherichia coli

Abstract A micro method for the isolation and characterization of the penicillin-binding sites in penicillin-binding proteins (PBPs) was developed. Only 10 nmol of a pure PBP are required for the whole procedure which is based on high-pressure liquid chromatography (HPLC). We showed that serine 44 in PBP 5 from Escherichia coli binds penicillin covalently.     

Seasonal succession of planktonic and epiphytic algae in a canal in Southern England

The standing crop of phytoplankton in a canal in southern England remained low during 1973 and 1974, seldom exceeding 5 × 10⁴ cells/1. Since phosphorus, nitrogen and silicon occurred abundantly in the water, competition with higher plants for some other substance must have limited development. Although the standing crop of epiphytic algae associated withNasturtium officinale andGroenlandia densa seemed to be limited by the number of attachment sites, this factor was of little importance in the case of algae attached toCladophora glomerata. Achnanthes minutissima v.cryptocephala was always predominant in the epiphytic assemblages, representing 50–8o% by numbers of the flora. The limited pool of predominant epiphytic taxa may have restricted the communities' ability to adapt to fluctuations in environmental conditions. The grazing of isopods, amphipods and molluscs probably never limited algal densities.     

In vitro construction of a recombinant adenovirus Ad2:Ad5

A hybrid virus containing the left half of the Ad5 genome and the right half of the Ad2 genome has been constructed by ligating together in vitro the BamHI.-A fragment of Ad5 (map co-ordinates 0–59.5) to the-SawHI-A fragment of Ad2 (map coordinates 59.5–100), and using this DNA to transfect susceptible cells. Viable progeny virus has been obtained which grows as well as the parental virus without any requirement for helper virus, and probably contains a hybrid hexon polypeptide consisting of the major part of the Ad5 hexon with an Ad2 carboxy terminus.     

Metal-directed affinity labeling of zinc(II), cobalt(II) and cadmium(II) horse liver alcohol dehydrogenases

The active site metal in horse liver alcohol dehydrogenase has been studied by metal-directed affinity labeling of the native zinc(II) enzyme and that substituted with cobalt(II) or cadmium(II). Reversible binding of bromoimidazolyl propionic acid to the cobalt enzyme blueshifts the visible absorption band originating from the catalytic cobalt atom at 655 to 630 nm. Binding of imidazole to the cobalt(II) enzyme redshifts the 655 nm band to 667 nm. Addition of bromoimidazolyl propionic acid blueshifts this 667 nm band back to 630 nm. This proves direct binding of the label to the active site metal in competition with imidazole. The affinity of the label for the reversible binding site in the three enzymes follows the order Zn ? Cd ? Co. After reversible complex formation, bromoimidazolyl propionic acid alkylates cysteine-46, one of the protein ligands to the active site metal. The nucleophilic reactivity of this metal-mercaptide bond in each reversible complex follows the order Co ? Zn ? Cd.     

Oviposition by the African migratory locust, Locusta migratoria migratorioides, in a crop environment in South Africa

Oviposition by the African migratory locust, Locusta migratoria migratorioides (Orthoptera: Acrididae), was studied in maize and wheat crops on the Orange Free State Highveld. Maize was shown to be the most important oviposition habitat with peak laying taking place in autumn and early winter when highest pod densities were recorded. Laying was mainly concentrated along the middle of the crop interrows in maize and within clearings in the wheat crop. Despite the uniform layout of these crops, the distribution of egg pods was found to be aggregated. Non-reproductive behaviour, such as locust aggregation, basking and feeding, as well as environmental factors appeared to influence the distribution of egg pods in these crops. Secondary selection for optinum soil moisture and compaction on the laying site enhanced the aggregation of pods.     

Contrasting Neurochemical Interactions of Tiletamine, a Potent Phencyclidine (PCP) Receptor Ligand, with the N-Methyl-D-Aspartate-Coupled and -Uncoupled PCP Recognition Sites

Neurochemical interactions of tiletamine, a potent phencyclidine (PCP) receptor ligand, with the N-methyl-D-aspartate (NMDA)-coupled and -uncoupled PCP recognition sites were examined. Tiletamine potently displaced the binding of [3H]1-(2-thienyl)cyclohexylpiperidine with an IC50 of 79 nM without affecting sigma-, glycine, glutamate, kainate, quisqualate, or dopamine (DA) receptors. Like other PCP ligands acting via the NMDA-coupled PCP recognition sites, tiletamine decreased basal, harmaline-, and D-serine-mediated increases in cyclic cGMP levels and induced stereotypy and ataxia. Tiletamine was nearly five times more potent than PCP at inhibiting the binding of 3-hydroxy[3H]PCP to its high-affinity NMDA-uncoupled PCP recognition sites. However, following parenteral administration, dizocilpine maleate (MK-801), ketamine, PCP, dexoxadrol, and 1-(2-thienyl)cyclohexylpiperidine HCl, but not tiletamine, increased rat pyriform cortical DA metabolism and/or release, a response modulated by the NMDA-uncoupled PCP recognition sites. Pretreatment with tiletamine did not attenuate the MK-801-induced increases in rat pyriform cortical DA metabolism, a result suggesting that tiletamine is not a partial agonist of the NMDA-uncoupled PCP recognition sites in this region. However, following intracerebroventricular administration (100-500 micrograms/rat), tiletamine increased pyriform cortical DA metabolism with a bell-shaped dose-response curve. These data indicate a differential interaction of tiletamine with the NMDA-coupled and -uncoupled PCP recognition sites. The paradoxical effects of tiletamine suggest that tiletamine might activate receptor(s) or neuronal pathways of unknown pharmacology.