Solution structure of the N‐terminal domain of DC‐UbP/UBTD2 and its interaction with ubiquitin

DC‐UbP/UBTD2 is a ubiquitin (Ub) domain‐containing protein first identified from dendritic cells, and is implicated in ubiquitination pathway. The solution structure and backbone dynamics of the C‐terminal Ub‐like (UbL) domain were elucidated in our previous work. To further understand the biological function of DC‐UbP, we then solved the solution structure of the N‐terminal domain of DC‐UbP (DC‐UbP_N) and studied its Ub binding properties by NMR techniques. The results show that DC‐UbP_N holds a novel structural fold and acts as a Ub‐binding domain (UBD) but with low affinity. This implies that the DC‐UbP protein, composing of a combination of both UbL and UBD domains, might play an important role in regulating protein ubiquitination and delivery of ubiquitinated substrates in eukaryotic cells.     

Adaptation of human left ventricular volumes to the onset of supine exercise

The purpose of this study was to measure the changes and rates of adaptation of left ventricular volumes at the onset of exercise. Eight asymptomatic subjects, in whom intramyocardial markers had been implanted 3-6 years previously during aortocoronary bypass surgery, exercised in the supine position at a constant workload of 73.6 W for 5 min. Six also exercised first at 16.4 W, and then against a workload which progressively increased by 8.2 W every 15 s. Cardiac volumes were measured by computer assisted analysis of the motion of the implanted markers. In the constant workload test, cardiac output increased rapidly from 5.7 +/- 1 min-1 to 10.3 +/- 1.9 1 min-1 by 2 min and then increased more slowly to 10.8 +/- 2.0 1 min-1 by 5 min. The cardiac output increase was mainly due to an increase in heart rate from 68 +/- 12 beats min-1 to 120 +/- 16 beats min-1 with minimal changes in stroke volume. The time constant for the early increase in cardiac output was 45s and for heart rate, 35s. With progressively increasing workloads, there was an almost linear increase of heart rate and cardiac output, but these increased at a slower rate than during the early phase of the constant load exercise test. In conclusion: rapid changes in cardiac output during supine exercise were produced by changes in heart rate; changes in stroke volume provided minor adjustments to cardiac output; the end-diastolic volume was almost constant.     

Influence of Methanogenic Populations in Holocene Lacustrine Sediments Revealed by Clone Libraries and Fatty Acid Biogeochemistry

Methanogenic populations were investigated in subsaline Laguna Potrok Aike sediments, southern Argentina. Microbial density and activity were assessed via cell count and in situ ATP detection for the last ～11K years. Methanogen phylogenetics highlighted species stratification throughout depth, whereas CO₂ reduction was the major pathway leading to methane production. Organic substrates, characterized using pore water analysis, bulk organic fractions and saturated fatty acids, showed a clear link between sediment colonization and initial organic sources. Concentrations and δ¹³C compositions of methane and fatty acids provided final evidence of a microbial imprint on Holocene organic proxies in the most colonized intervals.     

Interaction between the Effects of Inside and Outside Na and K on Bullfrog Skin Potential

The composition of the solution bathing one border of the isolated frog skin affects the response of the potential across the skin to changes in the composition of the solution bathing the opposite border. Increasing the K concentration of the inside (corium) bathing solution decreased the sensitivity of the potential to a change in outside Na concentration. Decreasing the outside Na concentration decreased the sensitivity of the potential to a change in inside K concentration. Increasing the total ionic strength of the outside bathing solution or of both bathing solutions decreased the sensitivity of the potential to a change in outside Na concentration.     

Effect of the uvs-2 allele of Neurospora crassa on the mutagenic potency of two N-hydroxylaminopurines and 2-aminopurine in the ad-3 forward-mutation test

The mutagenic potencies of 3 purine analogs were determined in the ad-3 forward-mutation test in growing cultures of heterokaryon 59 (H-59), a nucleotide excision repair-deficient (uvs-2/uvs-2) 2-component heterokaryon of Neurospora crassa. Two N-hydroxylaminopurines, 2-amino-6-N-hydroxylaminopurine (AHA) and 6-N-hydroxylaminopurine (HAP), were potent and strong mutagens, respectively, whereas 2-aminopurine (AP) was a moderate mutagen. Dose-response curves showed that AHA and HAP were about equally mutagenic at low doses but that AHA was more mutagenic than HAP at high doses. Comparison of these results in H-59 with our earlier results in heterokaryon 12 (H-12) of N. crassa, which is identical to H-59 except for being DNA-repair-proficient (uvs-2+/uvs-2+), shows that the defect in nucleotide excision repair due to uvs-2 has little or no effect on the mutagenic potencies of these 3 purine analogs. Therefore, the nucleotide excision-repair pathway in N. crassa that is deficient in H-59 does not appear to have a major role in the repair of pre-mutational lesions induced by these 3 purine analogs. On the other hand, based on the controls of these experiments, the frequency of spontaneous ad-3 mutants was 4 greater in H-59 than in H-12. This result suggests that the nucleotide excision-repair pathway in N. crassa that is inactivated by the uvs-2 mutation has a major role in the repair of lesions that would lead to spontaneous mutation at the ad-3+ region if they were not repaired.     

Isolation of heparin-binding growth factors from bovine, porcine and canine hearts

Fresh bovine, porcine and canine hearts were homogenized and mitogens for mesoderm-derived cells were purified in three different steps. Extraction by two different ammonium sulfate precipitations was followed by cation-exchange chromatography and by heparin-Sepharose affinity chromatography. A heparin-Sepharose fraction from heart (eluted at 1.1 M NaCl) increased mitotic activity in serum-deprived cultures of porcine aortic endothelial and smooth muscle cells, and in human fibroblasts. This mitogenic activity is potentiated by heparin and inhibited by gamma-interferon. The heart mitogenic fraction showed one double peak on HPLC at A215 and one polypeptide band on SDS/PAGE. These peaks and bands were identical to those obtained from bovine brain. The heart acidic fibroblast growth factor (aFGF) showed a positive signal in Western blots using antibodies raised against brain aFGF. Gas-phase amino acid sequencing established that the mitogens were identical to aFGF and the N-terminally truncated aFGF. Extraction in the presence of a protease inhibitor (pepstatin A) produced a higher-molecular mass form of aFGF with a blocked amino terminus. Another mitogen, eluted at 1.6 M NaCl from heparin-Sepharose, reacted with polyclonal antiserum against human recombinant basic fibroblast growth factor (bFGF) and showed a 66% (12 from 18 amino acids determined by gas-phase sequencing) similarity with bFGF. This polypeptide increased the mitotic activity of the same cell lines but was more potent than aFGF.