Conformational Studies of PACAP(1–27) and Its Fragments

Summary Conformational features of the neuropeptide pituitary adenylate cyclase activating polypeptide (1–27) (PACAP(1–27)) and its shorter fragments (1–5), (7–11) and (14–27) were studied by circular dichroism (CD) and fluorescence spectroscopy. The obtained CD spectra revealed that only PACAP(1–27) and the fragment (14–27) possess some content of an organized structure — the α-helix. This C-terminal, helical part of the peptides is important for receptor binding as it provides a stable structure that can reside in the ordered lipid region of the receptor site in the membrane, while the primary biological function of the hormone resides in the N-terminal, disordered part. Fluorescence studies have revealed that the tyrosine residue located in the helical region of PACAP has a higher quantum yield and a longer average lifetime than the tyrosine in the N-terminus, probably due to a ‘shielding’ effect of the hydrophobic cluster around Tyr²².     

Classification and prediction of clinical Alzheimer's diagnosis based on plasma signaling proteins

A molecular test for Alzheimer's disease could lead to better treatment and therapies. We found 18 signaling proteins in blood plasma that can be used to classify blinded samples from Alzheimer's and control subjects with close to 90% accuracy and to identify patients who had mild cognitive impairment that progressed to Alzheimer's disease 2-6 years later. Biological analysis of the 18 proteins points to systemic dysregulation of hematopoiesis, immune responses, apoptosis and neuronal support in presymptomatic Alzheimer's disease.     

Structural characterization of the O-specific polysaccharide from the lipopolysaccharide of the fish pathogen Aeromonas bestiarum strain P1S

The O-specific polysaccharide obtained by mild-acid degradation of lipopolysaccharide of Aeromonas bestiarum P1S was studied by sugar and methylation analyses along with ¹H and ¹³C NMR spectroscopy. The sequence of the sugar residues was determined using ¹H,¹H NOESY and ¹H,¹³C HMBC experiments. The O-specific polysaccharide was found to be a high-molecular-mass polysaccharide composed of tetrasaccharide repeating units of the structureSince small amounts of a terminal Quip3N residue were identified in methylation analysis, it was assumed that the elucidated structure also represented the biological repeating unit of the O-specific polysaccharide.     

Computer Modeling of the Solution Conformation of Cyclic Enkephalins

Summary The probable conformations of two cyclic enkephalin analogs, DNS-cyclo[d-Dab-Gly-Trp-Leu] (I) and DNS-cyclo[d-Dab-Gly-Trp-d-Leu] (II) (DNS=dansyl), were determined by combining the results of NOE, vicinal coupling constant and fluorescence energy transfer measurements with theoretical calculations. The common feature of the conformations for both peptides is the presence of a β-turn at residues 2 and 3.     

Local and long-range structural effects caused by the removal of the N-terminal polypeptide fragment from immunoglobulin L chain lambda

The role of the N-terminal polypeptide fragment of the immunoglobulin l-chain in V domain packing stability, and the flexibility of the whole chain was approached by molecular dynamics simulation. The observations were supported by experimental analysis. The N-terminal polypeptide fragment appeared to be the low-stability packing element in the V domain. At moderately elevated temperature it may be replaced at its packing locus by Congo red and then removed by proteolysis. After removal of Congo red by adsorption to (diethylamino)ethyl (DEAE) cellulose, the stability of complete L chain and of L chain devoid of the N-terminal polypeptide fragment were compared. The results indicated that the N-terminal polypeptide fragment plays an essential role in the stability of the V domain. Its removal makes the domain accessible for ANS and Congo red dye binding without heating. The decreased domain stability was registered in particular as increased root mean square (RMS) fluctuation and higher susceptibility to proteolytic attack. The long-range effect was most clearly manifested at 340 K as independent V and C domain fluctuation in the l-chain devoid of the N-terminal polypeptide fragment. This is likely due to the lack of direct connections between the N- and C-termini of the V domain polypeptide. In a complete V domain the connection involves residues 8-12 and 106-110 in particular. Partial or complete disruption of this connection increases the freedom of V domain rotation, while its increased cohesion strengthens the coupling of the V and C domains, making the whole L chain less flexible.     

Immunoexpression of androgen receptors and aromatase in testes of patient with Klinefelter's syndrome

Klinefelter's syndrome (47, XXY) is the most common chromosome aneuploidy in men and is usually characterized by underdeveloped testes and sterility. The aim of the present study was to detect cellular distribution of androgen receptors (AR) and aromatase in testes of patient with KS. The tissue sections were processed for morphological and immunohistochemical staining. Additionally, levels of FSH, LH, PRL, estradiol, and testosterone were measured in the plasma. Morphological analysis revealed a complete absence of spermatogenesis. No germ cells were present in seminiferous tubules. In some tubules, nests of apparently degenerating Sertoli cells were found. In the interstitium, Leydig cell hyperplasia was observed. Using immunohistochemistry, nuclear AR staining was detected in Sertoli cells and peritubular cells, whereas in Leydig cells the staining was exclusively cytoplasmic. The immunostaining of aromatase was detected in the cytoplasm of Sertoli cells and Leydig cells. Increased levels of gonadotropins and decreased level of testosterone concomitantly with the cytoplasmic localization of AR in Leydig cells might contribute to the impaired testicular function in patient with KS.     

Human herpesvirus 1 UL24 gene encodes a potential PD-(D/E)XK endonuclease

Using Meta-BASIC, a highly sensitive method for detection of distant similarity between proteins, we have identified another potential PD-(D/E)XK endonuclease in human herpesvirus 1 (HHV-1) encoded by the UL24 gene. The universal presence of UL24 in completed herpesviral genomes of three major subfamilies, Alphaherpesvirinae, Betaherpesvirinae, and Gammaherpesvirinae, suggests a fundamental role for this predicted PD-(D/E)XK endonuclease activity in the viral life cycle.     

Interactive effects of phosphate deficiency, sucrose and light/dark conditions on gene expression of UDP-glucose pyrophosphorylase in Arabidopsis

The effects of inorganic phosphate (Pi) status, light/dark and sucrose on expression of UDP-glucose pyrophosphorylase (UGPase) gene (Ugp), which is involved in sucrose/ polysaccharides metabolism, were investigated using Arabidopsis wild-type (wt) plants and mutants impaired in Pi and carbohydrate status. Generally, P-deficiency resulted in increased Ugp expression and enhanced UGPase activity and protein content, as found for wt plants grown on P-deficient and complete nutrient solution, as well as for pho1 (P-deficient) mutants. Ugp was highly expressed in darkened leaves of pho1, but not wt plants; daily light exposure enhanced Ugp expression both in wt and pho mutants. The pho1 and pho2 (Pi-accumulating) mutations had little or no effect on leaf contents of glucose and fructose, regardless of light/dark conditions, whereas pho1 plants had much higher levels of sucrose and starch in the dark than pho2 and wt plants. The Ugp was up-regulated when leaves were fed with sucrose in wt plants, but the expression in pho2 background was much less sensitive to sucrose supply than in wt and pho1 plants. Expression of Ugp in pgm1 and sex1 mutants (impaired in starch/sugar content) was not dependent on starch content, and not tightly correlated with soluble sugar status. Okadaic acid (OKA) effectively blocked the P-starvation and sucrose-dependent expression of Ugp in excised leaves, whereas staurosporine (STA) had only a small effect on both processes (especially in -P leaves), suggesting that P-starvation and sucrose effects on Ugp are transmitted by pathways that may share similar components with respect to their (in) sensitivity to OKA and STA. The results of this study suggest that Ugp expression is modulated by an interaction of signals derived from P-deficiency status, sucrose content and dark/light conditions, and that light/sucrose and P-deficiency may have additive effects on Ugp expression.     

Pharmacological doses of triiodothyronine upregulate lipogenic enzyme gene expression in rat white adipose tissue.

Triiodothyronine (T (3)) is known to increase liver lipogenic enzyme gene expression both in vivo and in tissue culture. Conflicting results have been reported on the effect of T (3) on lipogenic enzyme gene expression in white adipose tissue. The results presented in this paper indicate that administration of pharmacological doses of T (3) in rats leads to increased fatty acid synthase (FAS), acetyl-CoA carboxylase (ACC), ATP-citrate lyase (ACL) and malic enzyme (ME) activity in white adipose tissue. The increase in lipogenic enzyme activity was associated with increased FAS, ACC, ACL and ME mRNA levels. The response was dose-dependent. Activity of lipogenic enzyme and the lipogenic enzyme mRNA levels were positively correlated to serum T (3) concentration. The in vivo effect of T (3) on lipogenic enzyme gene expression could be reproduced in primary white rat adipocyte culture. In conclusion, the results presented in this paper indicate that T (3) exerts a stimulatory effect on lipogenic enzyme gene expression in white adipose tissue both in vivo and in tissue culture. Significant effects of T (3) on lipogenic enzyme gene expression were only observed in the presence of relatively high (pharmacological) concentrations of the hormone.