Amphiphilic spectroscopic probes utilizing metal chelates

Summary The synthesis and initial applications are reported for 1-[p-(pamitamido)-phenyl]ethylenedinitrilotetraacetic acid. The results demonstrate the versatility of this spectroscopic probe molecule, which allows achoice of a particular technique for a particular system as well as use of multiple spectroscopic techniques for complementary information about hydrophobic region in biological systems.This work was supported in part by grant No. CA-16861 from the National Cancer Institute, DHEW.     

Direct observation of enzyme substrate complexes by stopped-flow fluorescence: mathematical analyses

The fluorescence changes which occur upon the interaction of enzyme and substrate under stopped-flow conditions can provide a sensitive means to directly observe ES complexes. The interconversion of the intermediates during catalysis causes changes in fluorescence, signaling directly their existence, and allowing their quantitation. We have studied extensively an approach which measures radiationless energy transfer (RET) between enzyme tryptophanyl residues and a fluorescent peptide or ester substrate. Our studies of a number of proteolytic enzymes have validated the approach, which is sensitive and applicable to a variety of enzymes under a wide range of experimental conditions, including subzero temperatures. Direct excitation of fluorescent substrates can also be used to observe ES complex formation and breakdown and is complementary to the RET approach. Here we review both the RET and direct excitation kinetic approaches, with particular emphasis on the mathematical foundations we have developed which are critical to the successful interpretation of these or any other spectroscopic approach which yields a signal that is unique to the ES complex.     

Anomalous stability of insulin at very high pressure

Insulin is unaffected by pressures of 48,000 b in the solid state at room temperature, as elucidated both by spectroscopic measurements and bioassay. Its compression curve is reversible. The presence of water does not alter this property. Of a number of other proteins investigated insulin appears to be unique with regard to its pressure stability. The relative rigidity of the molecule combined with its small size may account for some of these properties.     

The renaissance of myoglobin: Dynamics,structure and oxygen binding control

Myoglobin as a prototype of a globular protein has been widely employed in advanced studies on the dynamics, folding and evolution of proteins. The advent of fast and super-fast lasers, the expression of site-directed mutants, and the application of advanced analytical and spectroscopic techniques have provided new insights into the structure-function relationships in proteins.     

Direct observation of enzyme substrate complexes by stopped-flow fluorescence: Mathematical analyses

Summary The fluorescence changes which occur upon the interaction of enzyme and substrate under stopped-flow conditions can provide a sensitive means to directly observe ES complexes. The interconversion of the intermediates during catalysis causes changes in fluorescence, signaling directly their existence, and allowing their quantitation. We have studied extensively and approach which measures radiationless energy transfer (RET) between enzyme tryptophanyl residues and a fluorescent peptide or ester substrate. Our studies of a number of proteolytic enzymes have validated the approach, which is sensitive and applicable to a variety of enzymes under a wide range of experimental conditions, including subzero temperatures. Direct excitation of fluorescent substrates can also be used to observe ES complex formation and breakdown and is complementary to the RET approach. Here we review both the RET and direct excitation kinetic approaches, with particular emphasis on the mathematical foundations we have developed which are critical to the successful interpretation of these or any other spectroscopic approach which yields a signal that is unique to ES complex.Acknowledgment. This work was supported by Grants-in-Aid GM-24967 and 24968 from the National Institute of Health of the Department of Health, Education and Welfare to Harvard Medical School. The continued advice and support of Dr B.L. Vallee is gratefully acknowledged, as is the help of Dr L. Bethune on theoretical aspects of the steady-state assumption. The excellent technical assistance of P. Maiorana is acknowledged. The abbreviations used are: RET; radiationless energy transfer, Dns, or dansyl; 5-dimethylaminonaphthalene-1-sulfonyl; DED; dansylethylenediamine, Mes; 2-(N-morpholino)ethane sulfonic acid, OMe; methoxide, mansyl; 6-(N-methylanilino)-naphthalene-2-sulfonyl.     

The isolation,identification and synthesis of the alarm pheromone ofVespula squamosa (Drury) (Hymenoptera: Vespidae) and associated behavior

Summary A material that elicits alarm and attack behavior byVespula squamosa (Drury) workers was isolated from venom extracts and identified by spectroscopic methods as N-3-methylbutylacetamide. This compound elicited attack responses from worker wasps identical to those responses observed when venom was applied at the same dosage. This is the first behavioral role reported for this compound.The authors thank E. Adamak, R. Murphy and F. Takken for technical assistance. This article reports the results of research only. Mention of a proprietary product does not constitute an endorsement or the recommendation for its use by USDA.     

Anomalous stability of insulin at very high pressure

Summary Insulin is unaffected by pressures of 48,000 b in the solid state at room temperature, as elucidated both by spectroscopic measurements and bioassay. Its compression curve is reversible. The presence of water does not alter this property. Of a number of other proteins investigated insulin appears to be unique with regard to its pressure stability. The relative rigidity of the molecule combined with its small size may account for some of these properties.Acknowlegments. I should like to thank Prof. Willard F. Libby of the Department of Chemistry and Institute of Geophysics of the University of California, Los Angeles, for his interest and support of this study.     

Molluscicidal and insecticidal activities of isobutylamides isolated fromFagara macrophylla

Summary Five isobutylamides were isolated as insect growth inhibitors and toxicants fromFagara macrophylla and identified from their spectroscopic data.Insects were kindly supplied by the agencies of the USDA in Tifton, GA; Phoenix, AZ and Brownsville TX. The authors thank Professor D. Heyneman forB. glabratus supply. This work was partly supported by a PHS Biomedical Grant to I.K.     

In vitro secretion of ecdysteroids by Y-organs of the crayfish,Procambarus clarkii

It was demonstrated that excised Y-organs of the crayfish,Procambarus clarkii, synthesize in vitro 3-dehydroecdysone (3-DHE) as the major product, together with small amounts of ecdysone. Both were identified by immunological and spectroscopic methods. The increase of ecdysteroidogenesis in the Y-organs was accompanied by an increase of the major free ecdysteroid, 20-hydroxyecdysone, in the hemolymph. This suggests a physiological role of 3-DHE, the details of which are still to be elucidated.     

Marine indoles of novel substitution pattern from the acorn wormGlossobalanus sp.

Summary 4,6-Dibromoindole and 4,6-dibromo-2-methylindole have been isolated from the acorn wormGlossobalanus sp. The biosynthetic implications of this finding are discussed.We wish to thank Prof. Paul J. Scheuer, University of Hawaii for his encouragement and for use of his facilities during spectroscopic work. T.H. gratefully acknowledges the giving of a travel grant by the University of the Ryukyus Foundation for International Exchange.     

Contribution to knowledge of the biosynthesis of cyclosporin A

3H- and 13C-NMR spectroscopic investigations on the structure of labeled cyclosporin A were performed after feeding of appropriate precursors. The 6 N-methyl groups and the methyl group in position 4 of the epsilon, zeta-unsaturated amino acid No. 1 (Mebmt) are introduced as intact CH3-units from methionine. Four head-to-tail acetate units are involved in the biosynthesis of the 8-carbon chain of the olefinic amino acid.     

Structure and stereochemistry of coccuvine (Cocculus laurifolius DC)

Summary Coccuvine C₁₇H₁₉NO₂, m.p. 137–38°, a new alkaloid fromCocculus laurifolius DC has been assigned the structure and stereochemistry as (I) on the basis of spectroscopic studies and chemical correlation.     

Roquefortine and isofumigaclavine A,metabolites fromPenicillium roqueforti

Summary The structures of two metabolites fromPenicillium roqueforti, designated roquefortine and isofumigaclavine A, have been determined by chemical and spectroscopic studies.     

Lanthanide-binding peptides and the enzymes that Might Have Been

The trivalent lanthanide ions are chemically similar to Ca(II) ions, making them useful Ca analogs for a multitude of applications. In addition, Ln(III) ions are efficient catalysts of hydrolysis due to their much stronger Lewis acidity relative to Ca(II) ions. Ln-binding peptides thus offer both the opportunity to study known Ca sites as well as to explore new biological functions with an entire family of spectroscopically rich and reactive ions. This review discusses Ln-binding peptides in three roles: (i) as models of Ca-protein structure and function, (ii) as spectroscopic tags for protein expression and characterization and (iii) as designed artificial endonucleases. The creation of hydrolytically active Ln peptides that can fold, bind, cleave and discriminate among substrates shows that the design of Ln enzymes can be accomplished, and they will serve as versatile biochemical tools to investigate protein folding, structure and nuclease function.Received 10 April 2004; received after revision 17 May 2004; accepted 28 May 2004     

Structural determinants of protein folding

The last several decades have seen an explosion of knowledge in the field of structural biology. With critical advances in spectroscopic techniques in examining structures of biomacromolecules, in maturation of molecular biology techniques, as well as vast improvements in computation prowess, protein structures are now being elucidated at an unprecedented rate. In spite of all the recent advances, the protein folding puzzle remains as one of the fundamental biochemical challenges. A facet to this empiric problem is the structural determinants of protein folding. What are the driving forces that pivot a polypeptide chain to a specific conformation amongst the vast conformation space? In this review, we shall discuss some of the structural determinants to protein folding that have been identified in the recent decades.     

A new sterol from the spongeHaliclona chilensis (Thiele)

Summary From the spongeHaliclona chilensis (Thiele) a minor keto-sterol was isolated and characterized as 24-keto-cholesta-5,25-dien-3-ol by means of spectroscopic (¹H and¹³C-FT NMR, MS) methods.     

Ca-antagonistic substance from soft coral of the genus Sarcophyton

A 14-membered ring diterpenoid named cembrane, possessing Ca-antagonistic action on the isolated rabbit aorta, has been isolated from a soft coral of the genus Sarcophyton, and its structure established by X-ray and spectroscopic data.     

The evidential significance of thought experiment in science

The most promising way to regard thought experiment is as a species of experiment, alongside concrete experiment. Of the authors who take this view, many portray thought experiment as possessing evidential significance intrinsically. In contrast, concrete experiment is nowadays most convincingly portrayed as acquiring evidential significance in a particular area of science at a particular time in consequence of the persuasive efforts of scientists. I argue that the claim that thought experiment possesses evidential significance intrinsically is contradicted by the history of science. Thought experiment, like concrete experiment, has evidential significance only where particular assumptions—such as the Galilean doctrine of phenomena—are taken to hold; under alternative premises, in themselves equally defensible, thought experiment is evidentially inert.     

Transition metals in an experimental tumour system

Summary In this report we present an analysis by atomic absorption spectrometry of some of the transition metals Fe, Zn, Cu, Mn, Co, and Ni in tissues and nuclear fractions in an experimental tumour system.Acknowledgments. The atomic absorption spectroscopic analysis was done at the Health Physics Division, Bhabha Atomic Research Centre, Bombay. The author are indebted to Dr S.D. Soman, Director, Health Physics Division for providing facilities for the said analysis.