Insufficiently dehydrated hydrogen bonds as determinants of protein interactions

The prediction of binding sites and the understanding of interfaces associated with protein complexation remains an open problem in molecular biophysics. This work shows that a crucial factor in predicting and rationalizing protein-protein interfaces can be inferred by assessing the extent of intramolecular desolvation of backbone hydrogen bonds in monomeric structures. Our statistical analysis of native structures shows that, in the majority of soluble proteins, most backbone hydrogen bonds are thoroughly wrapped intramolecularly by nonpolar groups except for a few ones. These latter underwrapped hydrogen bonds may be dramatically stabilized by removal of water. This fact implies that packing defects are "sticky" in a way that decisively contributes to determining the binding sites for proteins, as an examination of numerous complexes demonstrates.     

Incomplete modified fast in obese early pubertal girls leads to an increase in 24-hour growth hormone concentration and a lessening of the circadian pattern in leptin

We studied nutrition and GH in eight obese girls, aged 6-11 yr. Blood was sampled every 15 min for 24 h. A 48-h diet providing 25% of assumed caloric needs was imposed, with repeat sampling during the last 24 h. Six nonfasting lean girls were also studied, and their mean GH was 3 times that of the obese girls in the fed state (P = 0.024). Dieting increased mean GH by 60% (P = 0.0028). There was no difference in pulse number for either group, but total secretion for lean girls was 3.9 times greater than that in obese girls during the fed state. With dieting, obese girls increased their total GH secretion by 60% (P = 0.010), but maintained lower total secretion, approximately 40% that of lean girls (P = 0.014). Mean leptin in obese girls in the fed state was 6.2 times greater than mean leptin in lean girls (P = 0.0001), with higher concentrations at night (P < 0.05) and lowering of total mean leptin while dieting. We conclude that in early pubertal obese girls, short-term caloric restriction partially reverses the low GH state that is characteristic of obesity. The change is concomitant with a decrease in leptin and a lessening of circadian differences.     

Dramatic acceleration of protein folding by stabilization of a nonnative backbone conformation

Through a mutagenic investigation of Gly-48, a highly conserved position in the Src homology 3 domain, we have discovered a series of amino acid substitutions that are highly destabilizing, yet dramatically accelerate protein folding, some up to 10-fold compared with the wild-type rate. The unique folding properties of these mutants allowed for accurate measurement of their folding and unfolding rates in water with no denaturant by using an NMR spin relaxation dispersion technique. A strong correlation was found between beta-sheet propensity and the folding rates of the Gly-48 mutants, even though Gly-48 lies in an unusual non-beta-strand backbone conformation in the native state. This finding indicates that the accelerated folding rates of the Gly-48 mutants are the result of stabilization of a nonnative beta-strand conformation in the transition-state structure at this position, thus providing the first, to our knowledge, experimentally elucidated example of a mechanism by which folding can occur fastest through a nonnative conformation. We also demonstrate that residues that are most stabilizing in the transition-state structure are most destabilizing in the native state, and also cause the greatest reductions in in vitro functional activity. These data indicate that the unusual native conformation of the Gly-48 position is important for function, and that evolutionary selection for function can result in a domain that folds at a rate far below the maximum possible.     

Growth Hormone (GH) Secretion in Primary Adrenal Insufficiency: Effects of Cortisol Withdrawal and Patterned Replacement on GH Pulsatility and Circadian Rhythmicity

We studied the effects of cortisol withdrawal and patterned replacement upon spontaneous GH secretion and circadian rhythmicity in 7 patients with Addison's disease. Hydrocortisone was administered in physiological daily total dosages, and all resulting plasma cortisol values were 2–15 g/dl. It was given in 3 pulsatile modes: simulating physiological rhythm, reverse diurnal rhythmicity and continuous pulsatility. All modes of cortisol administration increased mean 24h, GH pulse amplitude and interpulse GH levels. During saline infusions circadian GH rhythmicity was preserved, with GH being at its highest between 2400–0400 h. Administration of hydrocortisone in any mode did not modify circadian GH rhythmicity. We conclude: Cortisol replacement in physiological daily doses increases GH output in patients with Addison's disease by augmenting GH pulse amplitude and interpulse levels. This is likely due to the attenuation of hypothalamic somatostatin (SRIF) secretion by physiologic levels of cortisol. By inference, it implies that cortisol deficiency leads to diminution of GH output with low GH pulse amplitude, likely as a result of an augmented hypothalamic somatostatin secretion. However, circadian rhythmicity of GH secretion is glucocorticoid-independent.     

Association of oestrogen receptor alpha gene polymorphism with the angiographic extent of coronary artery disease.

AIMS: To investigate the association between sequence variants in the promoter region of the oestrogen receptor-alpha (ER-alpha) gene and the angiographic severity of coronary artery disease (CAD). METHODS AND RESULTS: We studied 503 subjects undergoing coronary angiography (mean age 63+/-12 years, 72% men, 28% women). Coronary artery disease extent was assessed by the number of: (1) major coronary vessels with >50% narrowing (NMCV); (2) coronary vessels with any narrowing (NCV); (3) narrowed coronary segments (NCS). The number of thymine and adenine dinucleotide repeats [(TA)(n)], 1174 base-pairs upstream exon 1, was determined by PCR. The median number of (TA)(n)(18) was used to categorize subjects into long, short and mixed allele genotypes. Poisson regression was used to analyse the association between genotypes and CAD extent, with age category (age #10877;55 vs >55), sex, risk factors and age at onset of CAD as covariates. In young subjects, (TA)(n)length had a significant effect on NCS (P=0.047) and a borderline significant effect on NCV (P=0.066). Young subjects homozygous for long alleles had higher NCV and NCS compared to those homozygous for short alleles (NCV 3.7+/-2.4 vs 2.4+/-1.8, NCS 4.4+/-2.7 vs 3.1+/-2.3, respectively, P#10877;0.034). CONCLUSION: The (TA)(n)length in the ER-alpha gene promoter region is associated with the angiographic severity of CAD in young patients.     

The association of total cholesterol with processing speed is moderated by age in mid- to late-age healthy adults

Objectives: To investigate the nature of the association of normal levels of total cholesterol with cognitive function and the contribution of age to this association.

Methods: A sample of 61 senior executives, who were summoned for an annual medical examination with approximately four measurements of total cholesterol during 4 years, were examined with a computerized cognitive battery assessing mental processing speed as a sensitive measure of cognitive decline. We examined the association of total cholesterol with processing speed and the moderating effect of age on this association.

Results: A multiple regression analysis yielded a significant interaction between cholesterol and age for processing speed (p = .045). In order to examine the source of the interaction, simple slope analysis was performed. A significant negative high correlation was found for young subjects (p = .021), while no significant correlation was observed at middle (p = .286) or older (p = .584) age. The difference in slopes was robust to adjustment for potential confounding factors, including body mass index, and fasting glucose.

Conclusions: Within the normal range, higher total cholesterol levels were associated with better processing speed in younger ages and this association diminished with increasing age. Our findings highlight the important role of brain cholesterol in good cognitive functioning.     

OxNASH Score Correlates with Histologic Features and Severity of Nonalcoholic Fatty Liver Disease

Background and Aim

Oxidative stress is a core abnormality responsible for disease progression in nonalcoholic fatty liver disease (NAFLD). By employing a highly sensitive liquid chromatography–tandem mass spectrometry (LC/MS/MS) approach we recently were able to define the circulating profile of bioactive lipid peroxidation products characteristic of patients with nonalcoholic steatohepatitis (NASH) and developed the OxNASH score for NASH diagnosis. The aims of this study were to assess the utility of OxNASH as a predictor of NASH and study the association between OxNASH and specific histologic features of NAFLD.

Methods

Our cohort consisted of 122 patients undergoing liver biopsy for clinical suspicion of NAFLD. The NAFLD activity score (NAS) was calculated for each patient. Levels of fatty acid oxidation products were quantified using stable isotope dilution LC/MS/MS, and OxNASH was calculated.

Results

The mean age of our patients was 49.3 (±11.6) years, and the mean body mass index was 31.5 (±4.8) kg/m². The majority of patients were Caucasian (82 %) and 48 % were female. OxNASH correlated with NAS and with the individual histologic features of NAFLD, namely, steatosis, inflammation, and ballooning (P < 0.05), with the strongest association being with inflammation [rho (ρ) 0.40, 95 % confidence interval 0.23, 0.57, P < 0.001]. There was also a correlation between the stage of fibrosis and OxNASH (P = 0.001). These associations remained statistically significant after adjustment for multiple confounders.

Conclusions

Based on our results, in adult patients with NAFLD, OxNASH correlates with histologic features of NASH and appears to be a promising noninvasive marker.