Long-Acting Human Growth Hormone Analogue by Noncovalent Albumin Binding.

The present work describes a series of human growth hormone (hGH) albumin binder conjugates with an extended in vivo half-life. A broad range of different conjugates were studied by varying the albumin binder structure and conjugation site. Conjugates were conveniently obtained by reductive alkylation or by alkylation to introduced cysteines using functionalized albumin-binding side chains. In vitro and in vivo profiling provided the basis for identification of position L101C in human growth hormone as the most optimal position for conjugation, where both a sufficient level of receptor binding and a suitably long half-life could yield a molecule with potential for a once-weekly dosing regimen.

Systematic Review of Sexual Dysfunction Among Veterans with Post-Traumatic Stress Disorder.

INTRODUCTION: The clinical observations that many Vietnam veterans complained of sexual problems after returning from active duty have led to the question of a correlation between post-traumatic stress disorder (PTSD) and sexual dysfunction (SD). AIM: The purpose of this review is to systematically review the current literature regarding SD in male veterans with PTSD. METHODS: A systematic literature search, primarily in PubMed, the Cochrane database, and PsycINFO, was conducted. The keywords Sexual Dysfunction, Psychological OR Sexual Dysfunction, Physiological AND Stress Disorders, and Post-Traumatic were used. All manuscripts with relevance to the aim of the review were reviewed and considered. MAIN OUTCOME MEASURE: A total of 123 results were generated from the search. There were 11 publications regarding SD in veterans with PTSD included in the review. RESULTS: The included studies are described in detail in the Results section. All but one study found an increased and significant prevalence of SD among male veterans with PTSD, especially erectile dysfunction and decreased sexual desire. SD increased in patients with PTSD, with a prevalence between 8.4% and 88.6%; the large prevalence range were partly the result of methodological differences. Only two studies have examined the correlation between the severity of PTSD symptoms and SD, with conflicting results. Samples were of relatively moderate size. Only a few confounding factors were accounted for in the included studies. CONCLUSION: Increasing evidence suggests a correlation between PTSD and SD, but still, relatively few studies have addressed these questions. Further investigation is needed into the correlation between PTSD and SD, preferably taking severity of PTSD symptoms into account, along with confounders such as use of psychotropic medication, somatic illness, drug and alcohol abuse, and comorbid psychiatric illness. Bentsen IL, Giraldi AGE, Kristensen E, and Andersen HS. Systematic review of sexual dysfunction among veterans with post-traumatic stress disorder. Sex Med Rev 2015;3:78-87.

Tsunami-affected Scandinavian tourists: disaster exposure and post-traumatic stress symptoms.

BACKGROUND: Studies of short- and long-term mental effects of natural disasters have reported a high prevalence of post-traumatic stress. Less is known about disaster-exposed tourists repatriated to stable societies. AIMS: To examine the association between exposure to the 2004 Southeast Asian tsunami and symptoms of post-traumatic stress in three Scandinavian tourist populations. METHODS: Postal survey of Norwegian, Danish and Swedish Southeast Asia tourists registered by the police when arriving at national airports. Follow-up time was 6 (Norway), 10 (Denmark) and 14 months (Sweden) post-disaster; 6772 individuals were included and categorized according to disaster exposure: danger exposed (caught or chased by the waves), non-danger exposed (other disaster-related stressors) and non-exposed. Outcome measures were the Impact of Event Scale-Revised (IES-R) and Post Traumatic Stress Disorder Check List (PCL). RESULTS: Danger exposed reported more post-traumatic stress than non-danger exposed, and the latter reported more symptoms than non-exposed (each P<0.001). Comparison of the Norwegian and Swedish data suggested that symptoms were attenuated at 14 months follow-up (P<0.001). Female gender and low education, but not age, predicted higher levels of symptoms. CONCLUSIONS: Disaster-exposed tourists repatriated to unaffected home environments show long-term post-traumatic stress disorder symptoms related to the severity of exposure.

Residue 259 in protein-tyrosine phosphatase PTP1B and PTPalpha determines the flexibility of glutamine 262.

To study the flexibility of the substrate-binding site and in particular of Gln262, we have performed adiabatic conformational search and molecular dynamics simulations on the crystal structure of the catalytic domain of wild-type protein-tyrosine phosphatase (PTP) 1B, a mutant PTP1B(R47V,D48N,M258C,G259Q), and a model of the catalytically active form of PTPalpha. For each molecule two cases were modeled: the Michaelis-Menten complex with the substrate analogue p-nitrophenyl phosphate (p-PNPP) bound to the active site and the cysteine-phosphor complex, each corresponding to the first and second step of the phosphate hydrolysis. Analyses of the trajectories revealed that in the cysteine-phosphor complex of PTP1B, Gln262 oscillates freely between the bound phosphate group and Gly259 frequently forming, as observed in the crystal structure, a hydrogen bond with the backbone oxygen of Gly259. In contrast, the movement of Gln262 is restricted in PTPalpha and the mutant due to interactions with Gln259 reducing the frequency of the oscillation of Gln262 and thereby delaying the positioning of this residue for the second step in the catalysis, as reflected experimentally by a reduction in k(cat). Additionally, in the simulation with the Michaelis-Menten complexes, we found that a glutamine in position 259 induces steric hindrance by pushing the Gln262 side chain further toward the substrate and thereby negatively affecting K(m) as indicated by kinetic studies. Detailed analysis of the water structure around Gln262 and the active site Cys215 reveals that the probability of finding a water molecule correctly positioned for catalysis is much larger in PTP1B than in PTP1B(R47V,D48N,M258C,G259Q) and PTPalpha, in accordance with experiments.

Residue 182 influences the second step of protein-tyrosine phosphatase-mediated catalysis.

Previous enzyme kinetic and structural studies have revealed a critical role for Asp181 (PTP1B numbering) in PTP (protein-tyrosine phosphatase)-mediated catalysis. In the E-P (phosphoenzyme) formation step, Asp181 functions as a general acid, while in the E-P hydrolysis step it acts as a general base. Most of our understanding of the role of Asp181 is derived from studies with the Yersinia PTP and the mammalian PTP1B, and to some extent also TC (T-cell)-PTP and the related PTPa and PTPe. The neighbouring residue 182 is a phenylalanine in these four mammalian enzymes and a glutamine in Yersinia PTP. Surprisingly, little attention has been paid to the fact that this residue is a histidine in most other mammalian PTPs. Using a reciprocal single-point mutational approach with introduction of His182 in PTP1B and Phe182 in PTPH1, we demonstrate here that His182-PTPs, in comparison with Phe182-PTPs, have significantly decreased kcat values, and to a lesser degree, decreased kcat/Km values. Combined enzyme kinetic, X-ray crystallographic and molecular dynamics studies indicate that the effect of His182 is due to interactions with Asp181 and with Gln262. We conclude that residue 182 can modulate the functionality of both Asp181 and Gln262 and therefore affect the E-P hydrolysis step of PTP-mediated catalysis.

Imidazoline NNC77-0074 stimulates insulin secretion and inhibits glucagon release by control of Ca(2+)-dependent exocytosis in pancreatic alpha- and beta-cells.

We have investigated the effects of the novel imidazoline compound (+)-2-(2-(4,5-dihydro-1H-imidazol-2-yl)-thiopene-2-yl-ethyl)-pyridine (NNC77-0074) on stimulus-secretion coupling in isolated pancreatic alpha- and beta-cells. NNC77-0074 stimulated glucose-dependent insulin secretion in intact mouse pancreatic islets. No effect was observed at

Ligand-induced conformational changes: improved predictions of ligand binding conformations and affinities.

A computational docking strategy using multiple conformations of the target protein is discussed and evaluated. A series of low molecular weight, competitive, nonpeptide protein tyrosine phosphatase inhibitors are considered for which the x-ray crystallographic structures in complex with protein tyrosine phosphatase 1B (PTP1B) are known. To obtain a quantitative measure of the impact of conformational changes induced by the inhibitors, these were docked to the active site region of various structures of PTP1B using the docking program FlexX. Firstly, the inhibitors were docked to a PTP1B crystal structure cocrystallized with a hexapeptide. The estimated binding energies for various docking modes as well as the RMS differences between the docked compounds and the crystallographic structure were calculated. In this scenario the estimated binding energies were not predictive inasmuch as docking modes with low estimated binding energies corresponded to relatively large RMS differences when aligned with the corresponding crystal structure. Secondly, the inhibitors were docked to their parent protein structures in which they were cocrystallized. In this case, there was a good correlation between low predicted binding energy and a correct docking mode. Thirdly, to improve the predictability of the docking procedure in the general case, where only a single target protein structure is known, we evaluate an approach which takes possible protein side-chain conformational changes into account. Here, side chains exposed to the active site were considered in their allowed rotamer conformations and protein models containing all possible combinations of side-chain rotamers were generated. To evaluate which of these modeled active sites is the most likely binding site conformation for a certain inhibitor, the inhibitors were docked against all active site models. The receptor rotamer model corresponding to the lowest estimated binding energy is taken as the top candidate. Using this protocol, correct inhibitor binding modes could successfully be discriminated from proposed incorrect binding modes. Moreover, the ranking of the estimated ligand binding energies was in good agreement with experimentally observed binding affinities.