A 4D HCC(CO)NNH experiment for the correlation of aliphatic side-chain and backbone resonances in 13C/15N-labelled proteins

Summary We recently proposed a novel four-dimensional (4D) NMR strategy for the assignment of backbone nuclei in spectra of ¹³C/¹⁵N-labelled proteins (Boucher et al. (1992) J. Am. Chem. Soc., 114, 2262–2264 and J. Biomol. NMR, 2, 631–637). In this paper we extend this approach with a new constant time 4D HCC(CO)NNH experiment that also correlates the chemical shifts of the aliphatic sidechain (¹H and ¹³C) and backbone (¹H, ¹³C and ¹⁵N) nuclei. It separates the sidechain resonances, which may heavily overlap in spectra of proteins with large numbers of similar residues, according to the backbone nitrogen and amide proton chemical shifts. When used in conjunction with a 4D HCANNH or HNCAHA experiment it allows, in principle, complete assignment of aliphatic sidechain and backbone resonances with just two 4D NMR experiments.     

PHOTOSYNTHETIC PHYSIOLOGY OF PROROCENTRUM MARIAE-LEBOURIAE (DINOPHYCEAE) DURING ITS SUBPYCNOCLINE TRANSPORT IN CHESAPEAKE BAY1

This paper presents results of field studies on the estuarine dinoflagellate Prorocentrum mariae-lebouriae (Parke & Ballantine) Faust in Chesapeake Bay. We tested the hypothesis that the photosynthetic physiology of Prorocentrum shows adaptive responses to low-light during a lengthy subpycnocline transport in estuarine circulation. Prorocentrum underwent a seasonal, northward trnasport between February and June, 1984 and 1985. Low cell densities occurred in the seaward part of the estuary during winter and early-spring, subpycnocline populations progressed up-estuary in the ensuring 2–3 months, and dense surface populations developed in the mesohaline portion of the estuary thereafter. We sampled Prorocentrum from surface and subpycnocline waters and measured photosynthesis-light (P-I) relations with in situ incubations. The photophysiology of Prorocentrum collected below the pycnoline differed from that of cells in the surface mixed layer in that photosynthetic efficiency, α-cell^?1, was higher, photosynthetic capacity, P_max-cell^?1 was ·4 times greater for subpycnocline (≦ 10m) samples than for those from the surface mixed layer (≧ 6m). Comparison of in situ photosynthetic properties to those generated in laboratory studies showed that values of α·cell^?1 for both surface and subpycnocline samples were in the range found for cultures in low-light. Concentrations of Chls a, c and peridinin·cell^?1 and molar pigment ratios peridinin: Chl a and Chl a: Chl c were not significantly different for the surface and subpycnocline samples, nor were C · cell^?1 or C : Chl a. Chloroplast and starch volume fractions and the number of thylakoids were the same for samples collected at different depths, and there was no evidence of cytoplasmic vacuolization in any field samples. These morphometric data for cells from natural populations of Prorocentrum most closely resembled data for laboratory cultures grown at or near 2.6E·m-^?2·4d^?1. A lower growth irradiance of 0.3E·m^?2·d^?1 produced indications of stress in cultures, including starch depletion and vacuolization, that were never observed in natural populations. Based on the combination of these findings, we conclude that Prorocentrum is adapted to low-light both in the surface mixed layer and beneath the pycnocline, although certain photophysiological characteristics distinguish these two groups of samples.     

Morphogenetic movements during axolotl neural tube formation tracked by digital imaging

During neurulation in vertebrate embryos, epithelial cells of the neural plate undergo complex morphogenetic movements that culminate in rolling of the plate into a tube. Resolution of the determinants of this process requires an understanding of the precise movements of cells within the epithelial sheet. A computer algorithm that allows automated tracking of epithelial cells visible in digitized video images is presented. It is used to quantify the displacement field associated with morphogenetic movements in the axolotl (Ambystoma mexicanum) neural plate during normal neural tube formation. Movements from lateral to medial, axial elongations and area changes are calculated from the displacement field data and plotted as functions of time. Regional and temporal differences are identified. The approach presented is suitable for analyzing a wide variety of morphogenetic movements.     

Construction and analysis of a recombination-deficient (radA) mutant of Haloferax volcanii

By deleting the radA open reading frame of an extreme halophile, Haloferax volcanii , we created and characterized a recombination-deficient archaeon. This strain, Hf. volcanii DS52, has no detectable DNA recombination, is more sensitive to DNA damage by UV light and ethylmethane sulfonate, and has a slower growth rate than the wild type. These characteristics are similar to those observed in recombination mutants of Eukarya and Bacteria, and show that the radA gene belongs in the recA / RAD51 family by function as well as sequence homology. In addition, strain DS52 was not transformable by plasmids pWL102 or pUBP2 (which contain pHV2 and pHH1 replicons, respectively), although it was readily transformed by plasmids containing a pHK2 replicon, indicating a role for radA in the maintenance or replication of some halobacterial plasmids. Despite its slower growth rate, Hf. volcanii DS52 was still easy to culture and transform, and should be suitable for use in studies where a recombination-deficient background is desired.     

Weight loss and wrestling training: effects on growth-related hormones

Roemmich, James N., and Wayne E. Sinning. Weight lossand wrestling training: effects on growth-related hormones.J. Appl. Physiol. 82(6):1760-1764, 1997.Adolescent wrestlers(n = 9, 15.4 yr) and recreationallyactive control males (n = 7, 15.7 yr)were measured before, at the end of, and 3.5-4 mo after acompetitive wrestling season to assess the influence of dietary restriction on growth-related hormones. Wrestlers had significant elevations preseason to late season for morning serum concentrations (mean of 8 serial samples) of growth hormone (GH; 2.9 ± 0.7 vs. 6.5 ± 1.4 ng/ml) and sex hormone-binding globulin (SHBG; 16.1 ± 2.3 vs. 27.9 ± 6.9 nmol/l) and significant reductions in GH-binding protein (GHBP; 178 ± 19 vs. 109 ± 17 pmol/l), insulin-likegrowth factor I (IGF-I; 332 ± 30 vs. 267 ± 34 ng/ml),testosterone (T; 4.9 ± 0.4 vs. 3.6 ± 0.4 ng/ml), and freetestosterone (Free-T; 22.4 ± 3.6 vs. 15.7 ± 2.8 pg/ml).Wrestlers had significant postseason reductions in GH (3.44 ± 1.30 ng/ml) and SHBG (10.43 ± 4.13 nmol/l) but elevationsin GHBP (66.7 ± 23.8 pmol/l), IGF-I (72.9 ± 25.1 ng/ml),T (2.10 ± 0.46 ng/ml), and Free-T (9.76 ± 3.01 pg/ml). Concentrations of luteinizing hormone (LH), estradiol,prolactin, cortisol, insulin, and thyroid hormones did not differbecause of exercise-dietary practices of wrestlers. In-seasonelevations in GH, with concomitant reductions in GHBP and IGF-I, thatwere reversed during the postseason suggest a reduction in GH receptor number and partial GH resistance during the season. Nonelevated LH withreduced T levels suggests a central hypothalamic-pituitary-gonadal (H-P-G) axis impairment. In conclusion, undernutrition may lead toaltered H-P-G and GH-IGF-I axes function in adolescent wrestlers. However, only the wrestlers' late-season Free-T concentrations wereoutside the normal range, and the hormone axis impairments were quicklyreversed. The present data do not address hormonal axis responses toseveral years of wrestling and weight loss.