VOLUME 21, NO3, SEPT 1999

Signal transduction proteins: Structural basis of control by phosphorylation

Signal transduction proteins: Structural basis of control by phosphorylationread more

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N BROWN ,M E M NOBLE ,D BADFORD ,L N JOHNSON ,

Mechanism and action of myotoxin A from a rattlesnake venom

Mechanism and action of myotoxin A from a rattlesnake venomread more

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ANTHONY T TU ,

Blue copper proteins as honorary cytochromes: The structure and evolution of blue copper proteins

The blue copper proteins are among the most beautiful macromolecules that we know, and the intensity of the colour of azurin per copper atom is eighty times as that of the cuprammonium ion. They occur in the periplasm of Gram positive bacteria and in the photosystems of blue green algae (cyanobacteria) and in algal and higher plant chloroplasts. The function of these proteins in electron transport is very similar to those of the soluble cytochromes C-6 ("algal cytochrome f") and c(6) ("Pseudomonas cytochrome c-551"), so much so that one thinks of them as 'honorary cytochromes'. Several different sequence classes have been recognized, including azurin, plastocyanin, amicyanin, pseudoazurin and rusticyanin. There is considerable three-dimensional similarity in the copper binding sites of the proteins, and have as ligands two histidine, one cysteine and one methionine side chains. The proteins have been co-opted to function in a wide range of metabolisms, and azurin is often (but not always) associated with denitrification. The gene has been adapted to produce an outer membrane protein in gonococci, while azurin, amicyanin and pseudoazurin are involved in methylotrophy in different organisms - though in other closely;related bacteria, cytochromes c(8) play equally versatile roles. While the overall structures of each of the proteins are very similar wherever they are found, when duplicate genes occur in the same organism, the difference between the isoforms are large, even if no functional difference between the forms has been detected in vitro. The time is ripe and the techniques now available for the function and relationships of these proteins to be elucidated genetically, which has not been possible before because of the multiplicity of alternative pathways in bacterial electron transport systems.read more

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RICHARD P AMBER ,

Domain structure of the Rapana thomasiana (Gastropod) hemocyanin

Sufficient and well-dosed oxygen supply is one of the prerequisites of human and animal life and nature has developed families of respiratory proteins to master this task: Hemoglobins' myoglobins (vertebrates), hemocyanins (molluscs, arthropods) and hemerythrins (worms). Though structurally and in molecular size completely different, these proteins have all in common that under physiological conditions, oxygen is reversibly bound to a central heavy metal ion From these three families, the hemocyanins are one of the largest and most complex biopolymers, known so far, reaching diameters of up to 25 nm, comparable to ribosomes and detailed primary, secondary and quaternary structure determination obviously belongs to one of the most tedious tasks in biochemistry. The subunits of molluscan hemocyanins are organized into series of globular folded regions. Their structure has been clearly resolved in the electron microscope as a string of seven or eight beads (the number depending on the source of the hemocyanin), each presenting a functional oxygen-binding unit Between each pair of functional units is a short flexible linker region consisting of 10-15 amino acid residues. Rapana thomasiana hemocyanin aggregates are constituted of two different types of subunits called RHSS1 and RHSS2 which were purified by ion exchange chromatography of the dissociated Rapana hemocyanin Various tri-, di- and mono-domain fragments of the subunit are obtained by "autolysis" and limited proteolysis with V8 (Glu-C) proteinase, trypsin and elastase. The individual fragments are isolated by ion exchange, gel and/or reversed-phase chromatography. Their molecular weights are determined by SDS electrophoresis and the N-terminal amino acid sequences analyzed by Edman degradation, thus revealing the precise ordering of the domains in the Ropana subunit. The results confirm the 8-functional unit structure. Comparison of the RHSS1 domain sequences with those of other gastropod hemocyanin subunits is made.read more

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STANKA STOEVA ,WOLFGANG VOELTER ,PAVLINA DOLASHKA ,NICOLAY GENOV ,

Structure function studies of Clostridium perfringens alpha-toxin; A gas gangrene causing protein.

Structure function studies of Clostridium perfringens alpha-toxin; A gas gangrene causing protein.read more

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CLAIRE NAYLOR ,JULIAN EATON ,NEIL JUSTIN ,DAVID MOSS ,RICHARD TITBALL ,AJIT BASAK ,ANGELA HOWELLS ,

Elucidation of binding sites for protein denaturation by surfactant

One of the most important lessons that can be learned from studies of protein-surfactant interaction is that the details of the process reflect the detailed tertiary structures of the proteins. The binding of ionic surfactants to different water-soluble proteins; urease, peroxidase, human serum albumin and amino acid oxidase, were extensively studied by equilibrium dialysis and isothermal titration microcalorimetry techniques. The electrostatic interaction, which is accompanied by a preliminary hydrophobic interaction, occurs initially and is followed by a more extensive pure hydrophobic interaction The predominant unfolding of a protein is related to the first interaction, in which neutralization of charges at the surface of the protein perturb the balance of forces in the protein structure. The number of binding sited in the first set gl, is markedly consistent with concentration at midpoint of denaturation profiles and very close to the sites at the surface of the single subunit protein with opposite charge with respect to the ionic head group of surfactant.read more

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ALI A SABOURY ,ALI A MOOSAVI-MOVAHEDI ,

A genetic algorithm for low resolution protein structure determination

A genetic algorithm for iterative fitting of SAXS data is presented. The algorithm described produces fast convergence to a fittest model mass distribution compatible with experimental data. This method affords a dramatic reduction of processor time required by other SAXS fitting methods and can be applied to any kind of structure, the only requirement is the target profile. The effectiveness of the procedure is demonstrated with synthetic objects and by deriving the low resolution model of a known protein structure from their corresponding computed SAXS profile.read more

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J M ANDREU ,F MORAN ,J F DIAZ ,E PANTOS ,P CHACON ,

Structure-function relationships in sweet-tasting proteins

Six proteins have so far been isolated that to the higher primates. have an intensely sweet taste: monellin, thaumatin, brazzein, pentadin, mabinlin, and curculin. Curculin also acts as a taste-modifying protein, as does the protein miraculin. These proteins are all isolated from plants, but span a wide range of molecular weights and biochemical properties. Sequence comparisons show little similarity between the sweet proteins and suggest that they evolved independently of each other and are descendants of other proteins which wan originally parts of the molecular machinery of the plant. Structures haw been determined for several of the sweet proteins but these structures bear no resemblance to each other. Various method have been used to identify residues in these proteins that are responsible for their sweet taste. Thus far no common motif for a sweet determinant has been identified, although come immunological studies suggest that such a common structure may exist.read more

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JOHN L MARKLEY ,JANE CALDWELL ,

Recent studies on the abnormal hemoglobins found in Thailand

Numerous abnormal hemoglobins (Hb) have been discovered in Thailand, and yet further studies continue to show additional variants not previously described This paper reviews the discovery of two more variants, as well as additional studies on families with unusual combinations of mutations. One proposita had an abnormal Hb with similar electrophoretic mobility to Hb J Sangkok, but tryptic peptide mapping by h.p.l.c., amino acid analysis and protein sequence analysis revealed an abnormal peptide alpha-T9, due to the mutation [alpha 61 Lys-Asn], previously found in Hungary and identified as Hb J Buda. Direct DNA sequence analysis confirmed this result and showed that the mutation occurred in the alpha 1 gene and not the alpha 2-gene. The abnormal Hb in another proposita was shown by peptide mapping protein sequence analysis, and DNA sequence analysis, to be identical to Hb G Coushatta [beta 22 Glu-Ala]. In addition to the above variants, unusual combinations of mutations from two other families were studied namely the combination of Hb E (common in Thailand) with Hb C (common in African populations) and the combination of Hb beta(17)-thalassemia/ beta(41-42)-thalassemia.read more

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DALINA ITCHAYANAN ,PRANEE WIHICHAGOON ,WIPAPUN SIRIBOON ,CHANTRAGAN SRISOMSAP ,AREE LIMUTTIWONG ,JISNUSON SVASTI ,SUTHAT FUCHAROEN ,

Role of phosphatidylinositol 3-kinase in platelet aggregation

Platelet aggregation plays an important role in haemostasis and vascular disorders. This mainly takes place by the action of endogenous agonists like ADP, platelet-activating factor (PAF), epinephrine, 5-hydroxytryptamine (5-HT) and arachidonic acid (AA). These agonists except AA interact with G-protein coupled receptors Recent studies have shown that phosphatidylinositol 3-kinase (PI 3-kinase) and myosin light chain kinase (MLCK) play an important role in platelet aggregation We have shown that low doses of the selective PI 3-kinase inhibitor, wortmannin, inhibits aggregation (IC50; 20 nM) mediated by subthreshold doses of 5-hydroxytryptamine (5-HT; 5 mu M) and epinephrine (1 mu M). This study was undertaken to examine the effects of wortmannin in platelet aggregation induced by various platelet agonists. The results show that wortmannin inhibited aggregation mediated by various agonists with an IC50 for ADP (110 nM), AA (2 mu M), collagen (280 nM) and PAF (520 nhl). These studies suggest that wortmannin-mediated inhibition of aggregation induced by ADP, epinephrine, collagen and PAF may affect multiple enzymatic pathways and the most likely targets seem to be both PI 3-kinase and MLCK as other kinases like cAMP- and calcium-calmodulin-dependent protein kinases are reported to be not affected by wortmannin.read more

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S A SAEED ,Z NAWAZ ,B H SHAH ,G SHAMIM ,A H GILANI ,