Molecular heterogeneity of urinary proteins in wild house mouse populations.

Major urinary proteins (MUPs) from the urine of individual wild mice were characterized using electrospray ionization mass spectrometry (ESI-MS) and compared to MUPs from the urine of inbred mice. The wild mice showed considerable variation between individuals in the expression of a group of MUPs with similar masses. Some individuals excreted MUPs of unique molecular mass whilst some failed to express MUPs seen commonly in the other individuals. All the wild individuals contained proteins not previously observed in inbred mice. Urine from one individual was fractionated using anion exchange chromatography prior to analysis by ESI-MS. By analysing urine from inbred samples under the same conditions it was possible to relate, using mass and net charge in solution, MUPs from the wild sample to the MUPs that have been observed previously in inbred strains. This has allowed tentative identification of some MUPs from the wild mouse. The effect of collection history of urine from wild mice was also investigated. ESI-MS analysis of MUPs in a faecally contaminated sample showed the loss of a C-terminal tripeptide when compared to an uncontaminated sample from the same mouse, consistent with the presence of a specific endopeptidase. Similarly a sample of pooled urine provided by twelve individuals trapped from the same population showed evidence of loss of the C-terminal dipeptide.

First direct evidence for lipid/protein conjugation in oxidized human low density lipoprotein.

It has been postulated that lipids incorporated in atherosclerotic plaques are derived from the uptake of oxidized low density lipoprotein (LDL) by a macrophage-bound receptor. In vitro studies of LDL oxidation have established that reactive lipids are formed and that the exposure of native LDL to these products leads to modified protein with physical properties similar to oxidized LDL. Here we describe the application of highly specific tandem mass spectrometric techniques to the first characterization of lipid-modified LDL by demonstrating the addition of 4-hydroxy-2-nonenal to histidine residues of apolipoprotein B-100, following oxidation of LDL. The modified residues have been assigned to specific locations that have been previously shown to reside on the surface of the LDL particle.

Isolation and identification of lysophosphatidylcholines as endogenous modulators of thromboxane receptors.

Inhibition of thromboxane receptor radioligand binding to human platelet membranes has been employed as the basis for a radioreceptor assay designed to measure thromboxane receptor binding activity in samples of biological fluids. This method was used during phase 1 clinical evaluation of the thromboxane receptor antagonist SQ 30,741. Frequently, baseline plasma samples as well as plasma samples from placebo-treated subjects showed significant inhibition of radioligand binding in the radioreceptor assay, suggesting the presence of endogenous thromboxane receptor ligands. This receptor binding activity was stable and could be monitored in blood from normal volunteers using a modification of the radioreceptor assay. In order to identify the substance responsible for the observed activity, the activity present in pooled bovine blood was isolated and evaluated by a combination of FAB/MS, 1H-NMR, 13C-NMR and co-injection with reference standards on HPLC. Several endogenous thromboxane receptor ligands were identified as L-alpha-lysophosphatidylcholine (LPC) species. One major species, palmitoyl-LPC, contracted isolated rat aortic spirals, and these contractions could be delayed or prevented, but not reversed by the thromboxane receptor antagonist SQ 29,548. Palmitoyl-LPC slightly potentiated aortic contractions induced by the thromboxane receptor agonist, U-46,619, and diminished in a concentration-dependent manner the antagonism by SQ 29,548 of contractile responses to U-46,619. These findings are consistent with a potential for LPC species to bind and activate thromboxane receptors.

Dactylocyclines, novel tetracycline derivatives produced by a Dactylosporangium sp. II. Structure elucidation.

Fermentation of Dactylosporangium sp. (ATCC 53693) produces a mixture of tetracycline derivatives from which several related tetracycline glycosides, the dactylocyclines, were isolated and their structures determined. The most abundant glycoside in initial fermentations was found to be dactylocycline A. Each glycoside proved to be acid sensitive and readily hydrolyzed to a common aglycone, dactylocyclinone. While the aglycone was cross resistant with tetracycline, the dactylocyclines proved active against certain tetracycline-resistant organisms.

Lanomycin and glucolanomycin, antifungal agents produced by Pycnidiophora dispersa. II. Structure elucidation.

Two novel antifungal agents, lanomycin and glucolanomycin, as well as a biologically inactive degradation product, lanomycinol, were isolated from liquid fermentations of Pycnidiophora dispersa. All three compounds share an E,E,E-triene appended to a pyran ring. Lanomycin contains a glycine ester and glucolanomycin possesses a glucose unit attached to the glycine nitrogen. The structures, including absolute stereochemistry, were determined by spectroscopic analysis and partial chemical degradation. Both of the glycine containing compounds show activity against several pathogenic fungi in vitro.

Janthinocins A, B and C, novel peptide lactone antibiotics produced by Janthinobacterium lividum. II. Structure elucidation.

The structures of janthinocins A, B and C, three novel macrocyclic peptide lactone antibiotics isolated from fermentations of Janthinobacterium lividum, were determined. The janthinocins are of particular interest because they contain three amino acid residues that have not previously been reported in natural products: Each contains erythro-beta-hydroxy-D-leucine while janthinocins A and B also contain beta-hydroxytryptophan and beta-ketotryptophan, respectively.