Characterization of a substance P-Gly12 amidating enzyme in human cerebrospinal fluid

Enzyme activity capable of converting the glycine-extended substance P precursor, substance P-Gly12, into substance P was purified from human cerebrospinal fluid. The conversion reaction was monitored by radioimmunoassay measurement of substance P formation. The chemical identity of the product was verified by reversed-phase HPLC. The enzyme reaction was stimulated by Cu(II) ion and ascorbic acid and inhibited by the presence of diethyldithiocarbamate. By HPLC molecular sieving, the major enzyme activity appeared as a protein of 26,000 molecular weight.     

Hatchery selection promotes boldness in newly hatched brown trout (Salmo trutta): implications for dominance

By using newly hatched (approximately 2 weeks old) brown trout(Salmo trutta) from six families of wild and six families ofsea-ranched origin (seventh generation), we tested the hypothesesthat (1) the hatchery environment selects for increased boldness,and (2) boldness predicts dominance status. Sea-ranched troutspend their first 2 years in the hatchery before being releasedinto the wild at the onset of seaward migration. Trout werepresented with a novel object (tack) and with food (brine shrimp),and their responses were measured and scored in terms of boldness.Siblings with increasing difference in boldness were then pairedin dyadic contests. Fish of sea-ranged origin were on averagebolder than were fish of wild origin, and bolder individualswere more likely to become dominant regardless of origin. Boldnesswas not related to RNA levels, indicating that bold behaviorwas not a consequence of higher metabolism or growth rate. Neitherwas size a predictor of bold behavior or the outcome of dyadiccontests. These results are consistent with studies on olderlife stages showing increased boldness toward predators in hatchery-selectedfish, which suggests that behavioral consequences of hatcheryselection are manifested very early in life. The concordancebetween boldness and dominance may suggest that these behaviorsare linked in a risk prone-aggressive phenotype, which may bepromoted by hatchery selection. However, we also found significantvariation in behavioral and growth-related traits among families,suggesting that heritable variation has not been exhausted bysea-ranching procedures.     

A Treasure Chest of Balinese Images

Book reviewed in this article:
Images of Power: Balinese Paintings Made for Gregory Bateson and Margaret Mead . Hildred Geertz.     

Identification and design of a novel series of MGAT2 inhibitors

[Acyl CoA]monoacylglycerol acyltransferase 2 (MGAT2) is of interest as a target for therapeutic treatment of diabetes, obesity and other diseases which together constitute the metabolic syndrome. In this Letter we report our discovery and optimisation of a novel series of MGAT2 inhibitors. The development of the SAR of the series and a detailed discussion around some key parameters monitored and addressed during the lead generation phase will be given. The in vivo results from an oral lipid tolerance test (OLTT) using the MGAT2 inhibitor (S)-10, shows a significant reduction (68% inhibition relative to na?ve, p <0.01) in plasma triacylglycerol (TAG) concentration.     

Lipids and sterols of Pinus sylvestris L. sapwood and heartwood

Summary The lipid and sterol content and composition of three lipid fractions (free fatty acids/ sterols, triacylglycerols and sterol/triterpenoid esters) extracted from three stem discs of Pinus sylvestris were assessed to investigate metabolic changes related to heartwood formation. The wood was separated into (1) cambial zone, (2) outer sapwood, (3) inner sapwood, (4) transition zone, (5) outer heartwood and 6) inner heart-wood. The fractions were separated by thin-layer chromatography (TLC) and analysed by gas-liquid chromatography (GLC). The amount of fatty acids of sapwood triacylglycerols was about 1.5% (dry wt.) but a large reduction occurred in the transition zone. In contrast, noticeable amounts of free fatty acids were present only in the heart-wood. The most important fatty acids in the sapwood fractions were 16:0, 18:0, 18:1, 18:2 (the dominant fatty acid in all fractions), 18:3 and 20:3. Together 18:1 and 18:2 formed about 70% of the total triacylglycerol fatty acids. Of the sterol/ triterpenoid esters, 18:2 and 18:3 were predominant. The fatty acid composition of all fractions changed in the transition zone. The sterols found were sitosterol, stigmastanol, campesterol and campestanol. The amount of sterol esters increased towards the heartwood, and the amount of free sterols was lowest in the inner sapwood. Sitosterol was the dominant sterol in both free sterols and sterol esters.     

Energetics of complementary side-chain packing in a protein hydrophobic core

The energetics of complementary packing of nonpolar side chains in the hydrophobic core of a protein were analyzed by protein engineering experiments. We have made the mutations Ile----Val, Ile----Ala, and Leu----Ala in a region of the small bacterial ribonuclease barnase where the major alpha-helix packs onto the central beta-sheet. The destabilization resulting from the creation of cavities was determined by measuring the decrease in free energy of folding from reversible denaturation induced by urea, guanidinium chloride, or heat. The different methods give consistent and reproducible results. The loss in free energy of folding for the mutant proteins is 1.0-1.6 kcal/mol per methylene group removed. This exceeds by severalfold the values obtained from model experiments of the partitioning of relevant side chains between aqueous and nonpolar solvents. Much of this discrepancy arises because two surfaces are buried when a protein folds--both the amino acid side chain in question and the portions of the protein into which it packs. These experiments directly demonstrate that the interior packing of a protein is crucial in stabilizing its three-dimensional structure: the conversion of leucine or isoleucine to alanine in the hydrophobic core loses half the net free energy of folding of barnase with a concomitant decrease in yield of the expressed recombinant protein.     

Isoforms of a dynorphin B converting enzyme isolated by column electrophoresis in agarose suspension.

A dynorphin B converting enzyme previously purified from bovine spinal cord was subjected to column electrophoresis in agarose suspension. By this technique combined with HPLC gel filtration it was possible to resolve and recover several isoforms of the proteinase. All these isoenzymes were associated with a similar molecular size but apparently they differed with regard to their net charges. No significant difference between their inhibitory profiles or their Km values for the release of Leu-enkephalin-Arg6 from dynorphin B was observed.     

Molecular heterogeneity of angiotensin converting enzyme in human cerebrospinal fluid.

Three different molecular forms of angiotensin converting enzyme (ACE) (approximately Mr 150,000, 80,000 and 40,000, respectively), have been recovered from human cerebrospinal fluid. All three enzymes were inhibited by captopril and enalapril and their activity was potentiated by chloride ions. They were capable of degrading Leu-enkephalin-Arg6 and substance -P, but gave no conversion of neurokinin A. In all these aspects, the CSF enzymes were identical with the human pulmonary enzyme. The Mr 40,000 form of ACE is the smallest active form of the enzyme hitherto reported and is likely to represent a fragment of the C-terminal part of native ACE, where its active center is located.