Selective inhibition of proline hydroxylation by 3,4-dehydroproline

The effect of proline analogs on peptidyl proline hydroxylation has been studied in vivo using aerated root slices of Daucus carota. One analog, 3,4-dehydroproline, acted at micromolar concentrations to rapidly and selectively inhibit peptidyl proline hydroxylation. A structurally altered hydroxyproline-rich cell wall glycoprotein was synthesized and secreted by dehydroproline-treated tissue. The capacity to hydroxylate proline recovered slowly following a short pulse treatment with the analog, with a halftime for recovery of about 24 hours. Recovery was not altered by supplying exogenous proline. Dehydroproline had little effect on the induction of nitrate reductase by nitrate, nor on wound-induced increases in amino acid uptake and protein synthesis. In contrast, other proline analogs inhibit proline hydroxylation only at millimolar concentrations. It is hypothesized that dehydroproline acts as an enzyme-activated suicide inhibitor of prolyl hydroxylase. This analog should become a useful tool for elucidating the functional significance of hydroxyproline-rich glycoproteins.     

Disturbance and contrasting patterns of population structure in the brachiopod Terebratulina septentrionalis (Couthouy) from two subtidal habitats

The population structures of Terebratulina septentrionalis (Couthouy) from exposed upper rock surface and semi-cryptic rock wall habitats at 33 m depth in the Gulf of Maine differ. Over a 3-yr period, population densities were consistently higher in rock wall habitats. Although both populations were dominated by juveniles (1–4 mm shell length), size-frequency distributions constructed from upper rock surface and rock wall populations were significantly different, as a result of a greater frequency of large brachiopods (> 20 mm shell length) in rock wall populations. Prominent modes occurred at 14–15 mm shell length in upper surface populations and at 19–20 mm length in rock wall populations. Recruitment was higher in rock wall habitats where ambient light intensities were significantly lower than on upper rock surfaces. Differences in recruitment are either the result of larval selection for shaded rock walls or differential juvenile mortality between habitats. The larvae of Terebratulina settle on a diverse array of substrata. These include bedrock, sandy polychaete tubes and algae in upper surface habitats and bedrock, calcareous polychaete tubes, and ascidians in rock wall habitats. Individuals attached to polychaete tubes and algae in upper surface habitats do not attain large body size (> 13 mm shell length). It is suggested that these differences in population structure reflect the greater intensity of disturbance in upper surface habitats. For example, the cod, Gadus morhua (Linnaeus), ingests brachiopods attached to algae and polychaete tubes in this habitat. Gastropod predation affects brachiopods in upper surface habitats but not in rock wall habitats. Predation by gastropods and asteroids is not size-specific. These results are consistent with the hypothesis that predation contributed to the decline in the abundance and diversity of articulate brachiopods since the Mesozoic, and suggest that the restriction of recent populations to semi-cryptic rock wall and crevice habitats is, in part, controlled by disturbance.     

Altered gene products are associated with activation of cellular rasK genes in human lung and colon carcinomas

Two lung and two colon carcinoma cell lines of human origin, which contained the same activated ras^K transforming gene, expressed abnormal species of p21 that were distinct from the p21 proteins expressed in normal human cells and other human carcinomas. The abnormal species of p21 expressed by three of these cell lines were indistinguishable from each other, but differed from the abnormal p21 expressed by one lung carcinoma cell line. NIH cells transformed by DNAs of these carcinomas expressed the same abnormal p21 species, indicating that these abnormal proteins were encoded by the activated ras^K genes detected by transfection. These results indicate that transforming activity of ras^K genes in human lung and colon carcinoma cell lines is activated by mutations which alter the structure of their gene products, and that activation of ras^K genes can result from different molecular alterations in different individual neoplasms.     

Inhibition of neuronal sodium and potassium ion activated adenosinetriphosphatase by pyrithiamin

Since one of the electrophysiological effects of pyrithiamin, an antimetabolite of thiamin, suggested an interference with sodium pump mechanisms, the effect of pyrithiamin on Na+,K+-ATPase was investigated. We found that whereas preincubation of the antimetabolite with nonneuronal preparations of Na+,K+-ATPase produced only minimal inhibition, the enzyme derived from brain preparations was markedly inhibited. This inhibition could be prevented by thiamin but not reversed. The kinetic study showed that pyrithiamin acts in a noncompetitive manner with respect to the activation of the enzyme by ATP, Na+, and K+. Pyrithiamin inhibited Na+-dependent phosphorylation and K+-stimulated phosphatase as well as ouabain binding, and these inhibitions were parallel with that of the overall Na+,K+-ATPase reaction. In addition, the antimetabolite caused a significant change in the turbidity of the enzyme suspension. The results suggest that pyrithiamin may induce a structural change of the enzyme complex.     

Studies of the limited degradation of mucus glycoproteins. The effect of dilute hydrogen peroxide.

1. The action of dilute H2O2 on a series of ovarian-cyst glycoproteins and glycopolypeptides was investigated. 2. Both native glycoproteins and the glycopolypeptides were carbohydrate-rich, of relatively low molecular weight and of simple structure. 3. At pH 5.6 and 37 degrees C, exposure to H2O2 for a limited time brought about a partial degradation, the molecular weight being decreased by 2-4-fold. 4. Carbohydrate analysis showed very little change in the oligosaccharide moiety, apart from a small decrease in sialic acid in some samples. 5. Amino acid analysis showed minor changes in serine, threonine and proline contents, but almost total loss of histidine. Concomitantly, there was a small gain in aspartic acid. 6. Myosin, examined at both pH 5.7 and 6.7, exhibited generally similar behaviour, there being losses of other amino acid residues as well as histidine: the viscosity was decreased to a low value, and a range of peptides of widely varying size was produced. 7. It is suggested that attack on the histidine residue, with partial conversion into aspartic acid, is accompanied by scission of the histidyl peptide bond.     

Chicken lens delta-crystallin gene expression and methylation in several non-lens tissues.

RNA sequences coding for the most abundant chicken lens proteins, delta-crystallin, were detected at very low levels in day old post hatch chick lung, heart, kidney and liver, and in 6 day embryo headless bodies. The pattern of cytosine methylation within the CCGG sequences of the delta-crystallin genes was also examined and shown to vary in several non-lens tissues, from several stages of development. Embryonic neural retina, which expresses a higher level of delta-crystallin RNA than the above tissues, is no less methylated in the sites studied than the tissues which have no association with the eye, and is actually more heavily methylated than the kidney. Thus no obvious correlation was found between undermethylation and gene expression.     

Induction of the allantoin degradative enzymes by allophanic acid, the last intermediate of the pathway

$\text{Saccharomyces cerevisiae}$ can utilize allantoin as a sole nitrogen source by degrading it in five steps to ammonia, “CO₂”, and glyoxylate. We have previously shown that allophanic acid is the inducer of the urea carboxylase: allophanate hydrolase multienzyme complex. Since these enzymes catalyse the last two steps of allantoin degradation, experiments were performed to determine if allophanate was also the inducer of any other enzymes in the pathway. Our data demonstrate that allophanate induces synthesis of at least five of the seven purine degradative enzymes.