Scoring Protein Relationships in Functional Interaction Networks Predicted from Sequence Data

The abundance of diverse biological data from various sources constitutes a rich source of knowledge, which has the power to advance our understanding of organisms. This requires computational methods in order to integrate and exploit these data effectively and elucidate local and genome wide functional connections between protein pairs, thus enabling functional inferences for uncharacterized proteins. These biological data are primarily in the form of sequences, which determine functions, although functional properties of a protein can often be predicted from just the domains it contains. Thus, protein sequences and domains can be used to predict protein pair-wise functional relationships, and thus contribute to the function prediction process of uncharacterized proteins in order to ensure that knowledge is gained from sequencing efforts. In this work, we introduce information-theoretic based approaches to score protein-protein functional interaction pairs predicted from protein sequence similarity and conserved protein signature matches. The proposed schemes are effective for data-driven scoring of connections between protein pairs. We applied these schemes to the Mycobacterium tuberculosis proteome to produce a homology-based functional network of the organism with a high confidence and coverage. We use the network for predicting functions of uncharacterised proteins

Automated protein resonance assignments of magic angle spinning solid-state NMR spectra of β1 immunoglobulin binding domain of protein G (GB1)

Magic-angle spinning solid-state NMR (MAS SSNMR) represents a fast developing experimental technique with great potential to provide structural and dynamics information for proteins not amenable to other methods. However, few automated analysis tools are currently available for MAS SSNMR. We present a methodology for automating protein resonance assignments of MAS SSNMR spectral data and its application to experimental peak lists of the β1 immunoglobulin binding domain of protein G (GB1) derived from a uniformly 13C- and 15N-labeled sample. This application to the 56 amino acid GB1 produced an overall 84.1% assignment of the N, CO, CA, and CB resonances with no errors using peak lists from NCACX 3D, CANcoCA 3D, and CANCOCX 4D experiments. This proof of concept demonstrates the tractability of this problem

3D diffractive imaging of nanoparticle ensembles using an X-ray laser

We report the 3D structure determination of gold nanoparticles (AuNPs) by X-ray single particle imaging (SPI). Around 10 million diffraction patterns from gold nanoparticles were measured in less than 100 hours of beam time, more than 100 times the amount of data in any single prior SPI experiment, using the new capabilities of the European X-ray free electron laser which allow measurements of 1500 frames per second. A classification and structural sorting method was developed to disentangle the heterogeneity of the particles and to obtain a resolution of better than 3 nm. With these new experimental and analytical developments, we have entered a new era for the SPI method and the path towards close-to-atomic resolution imaging of biomolecules is apparent

British Society of Gastroenterology (BSG) and British Society of Paediatric Gastroenterology, Hepatology and Nutrition (BSPGHAN) joint consensus guidelines on the diagnosis and management of eosinophilic oesophagitis in children and adults

Background: Eosinophilic oesophagitis (EoE) is an increasingly common cause of dysphagia in both children and adults, as well as one of the most prevalent oesophageal diseases with a significant impact on physical health and quality of life. We have provided a single comprehensive guideline for both paediatric and adult gastroenterologists on current best practice for the evaluation and management of EoE. Methods: The Oesophageal Section of the British Society of Gastroenterology was commissioned by the Clinical Standards Service Committee to develop these guidelines. The Guideline Development Group included adult and paediatric gastroenterologists, surgeons, dietitians, allergists, pathologists and patient representatives. The Population, Intervention, Comparator and Outcomes process was used to generate questions for a systematic review of the evidence. Published evidence was reviewed and updated to June 2021. The Grading of Recommendations, Assessment, Development and Evaluation (GRADE) system was used to assess the evidence and make recommendations. Two rounds of voting were held to assess the level of agreement and the strength of recommendations, with 80% consensus required for acceptance. Results: Fifty-seven statements on EoE presentation, diagnosis, investigation, management and complications were produced with further statements created on areas for future research. Conclusions: These comprehensive adult and paediatric guidelines of the British Society of Gastroenterology and British Society of Paediatric Gastroenterology, Hepatology and Nutrition are based on evidence and expert consensus from a multidisciplinary group of healthcare professionals, including patient advocates and patient support groups, to help clinicians with the management patients with EoE and its complications

In vivo biofunctional evaluation of hydrogels for disc regeneration

Purpose Regenerative strategies aim to restore the original biofunctionality of the intervertebral disc. Different biomaterials are available, which might support disc regeneration. In the present study, the prospects of success of two hydrogels functionalized with anti-angiogenic peptides and seeded with bone marrow derived mononuclear cells (BMC), respectively, were investigated in an ovine nucleotomy model. Methods In a one-step procedure iliac crest aspirates were harvested and, subsequently, separated BMC were seeded on hydrogels and implanted into the ovine disc. For the cell-seeded approach a hyaluronic acid-based hydrogel was used. The anti-angiogenic potential of newly developed VEGF-blockers was investigated on ionically crosslinked metacrylated gellan gum hydrogels. Untreated discs served as nucleotomy controls. 24 adult merino sheep were used. After 6 weeks histological, after 12 weeks histological and biomechanical analyses were conducted. Results Biomechanical tests revealed no differences between any of the implanted and nucleotomized discs. All implanted discs significantly degenerated compared to intact discs. In contrast, there was no marked difference between implanted and nucleotomized discs. In tendency, albeit not significant, degeneration score and disc height index deteriorated for all but not for the cell-seeded hydrogels from 6 to 12 weeks. Cell-seeded hydrogels slightly decelerated degeneration. Conclusions None of the hydrogel configurations was able to regenerate biofunctionality of the intervertebral disc. This might presumably be caused by hydrogel extrusion. Great importance should be given to the development of annulus sealants, which effectively exploit the potential of (cell-seeded) hydrogels for biological disc regeneration and restoration of intervertebral disc functioningThis work was supported by the EU-project Disc Regeneration (NMP3-LA-2008-213904). Technical assistance of Iris Baum and the whole animal surgery team of the Institute of Orthopaedic Research and Biomechanics, Ulm, are gratefully acknowledged. DDAHA hydrogels were kindly provided by Cristina Longinotti (DDAHA, Anika Therapeutics, Abano Therme, Italy)

Phos-Tag-Based Analysis of Myosin Regulatory Light Chain Phosphorylation in Human Uterine Myocytes

The 'phosphate-binding tag' (phos-tag) reagent enables separation of phospho-proteins during SDS-PAGE by impeding migration proportional to their phosphorylation stoichiometry. Western blotting can then be used to detect and quantify the bands corresponding to the phospho-states of a target protein. We present a method for quantification of data regarding phospho-states derived from phos-tag SDS-PAGE. The method incorporates corrections for lane-to-lane loading variability and for the effects of drug vehicles thus enabling the comparison of multiple treatments by using the untreated cellular set-point as a reference. This method is exemplified by quantifying the phosphorylation of myosin regulatory light chain (RLC) in cultured human uterine myocytes.We have evaluated and validated the concept that, when using an antibody (Ab) against the total-protein, the sum of all phosphorylation states in a single lane represents a 'closed system' since all possible phospho-states and phosphoisotypes are detected. Using this approach, we demonstrate that oxytocin (OT) and calpeptin (Calp) induce RLC kinase (MLCK)- and rho-kinase (ROK)-dependent enhancements in phosphorylation of RLC at T18 and S19. Treatment of myocytes with a phorbol ester (PMA) induced phosphorylation of S1-RLC, which caused a mobility shift in the phos-tag matrices distinct from phosphorylation at S19.We have presented a method for analysis of phospho-state data that facilitates quantitative comparison to a reference control without the use of a traditional 'loading' or 'reference' standard. This analysis is useful for assessing effects of putative agonists and antagonists where all phospho-states are represented in control and experimental samples. We also demonstrated that phosphorylation of RLC at S1 is inducible in intact uterine myocytes, though the signal in the resting samples was not sufficiently abundant to allow quantification by the approach used here

Miiuy Croaker Hepcidin Gene and Comparative Analyses Reveal Evidence for Positive Selection

Hepcidin antimicrobial peptide (HAMP) is a small cysteine-rich peptide and a key molecule of the innate immune system against bacterial infections. Molecular cloning and genomic characterization of HAMP gene in the miiuy croaker (Miichthys miiuy) were reported in this study. The miiuy croaker HAMP was predicted to encode a prepropeptide of 99 amino acids, a tentative RX(K/R)R cleavage motif and eight characteristic cysteine residues were also identified. The gene organization is also similar to corresponding genes in mammals and fish consisting of three exons and two introns. Sequence polymorphism analysis showed that only two different sequences were identified and encoded two proteins in six individuals. As reported for most other species, the expression level was highest in liver and an up-regulation of transcription was seen in spleen, intestine and kidney examined at 24 h after injection of pathogenic bacteria, Vibrio anguillarum, the expression pattern implied that miiuy croaker HAMP is an important component of the first line defense against invading pathogens. In addition, we report on the underlying mechanism that maintains sequences diversity among fish and mammalian species, respectively. A series of site-model tests implemented in the CODEML program revealed that moderate positive Darwinian selection is likely to cause the molecular evolution in the fish HAMP2 genes and it also showed that the fish HAMP1 genes and HAMP2 genes under different selection pressures

Tension-Compression Loading with Chemical Stimulation Results in Additive Increases to Functional Properties of Anatomic Meniscal Constructs

Objective: This study aimed to improve the functional properties of anatomically-shaped meniscus constructs through simultaneous tension and compression mechanical stimulation in conjunction with chemical stimulation. Methods: Scaffoldless meniscal constructs were subjected to simultaneous tension and compressive stimulation and chemical stimulation. The temporal aspect of mechanical loadingwas studied by employing two separate five day stimulation periods. Chemical stimulation consisted of the application of a catabolic GAG-depleting enzyme, chondroitinase ABC (C-ABC), and an anabolic growth factor, TGF-b1. Mechanical and chemical stimulation combinations were studied through a full-factorial experimental design and assessed for histological, biochemical, and biomechanical properties following 4 wks of culture. Results: Mechanical loading applied from days 10–14 resulted in significant increases in compressive, tensile, and biochemical properties of meniscal constructs. When mechanical and chemical stimuliwere combined significant additive increases in collagen per wet weight (4-fold), compressive instantaneous (3-fold) and relaxation (2-fold) moduli, and tensile moduli in the circumferential (4-fold) and radial (6-fold) directions were obtained. Conclusions: This study demonstrates that a stimulation regimen of simultaneous tension and compression mechanical stimulation, C-ABC, and TGF-b1 is able to create anatomic meniscus constructs replicating the compressive mechanica

Serum Stabilities of Short Tryptophan- and Arginine-Rich Antimicrobial Peptide Analogs

Several short antimicrobial peptides that are rich in tryptophan and arginine residues were designed with a series of simple modifications such as end capping and cyclization. The two sets of hexapeptides are based on the Trp- and Arg-rich primary sequences from the "antimicrobial centre" of bovine lactoferricin as well as an antimicrobial sequence obtained through the screening of a hexapeptide combinatorial library.HPLC, mass spectrometry and antimicrobial assays were carried out to explore the consequences of the modifications on the serum stability and microbicidal activity of the peptides. The results show that C-terminal amidation increases the antimicrobial activity but that it makes little difference to its proteolytic degradation in human serum. On the other hand, N-terminal acetylation decreases the peptide activities but significantly increases their protease resistance. Peptide cyclization of the hexameric peptides was found to be highly effective for both serum stability and antimicrobial activity. However the two cyclization strategies employed have different effects, with disulfide cyclization resulting in more active peptides while backbone cyclization results in more proteolytically stable peptides. However, the benefit of backbone cyclization did not extend to longer 11-mer peptides derived from the same region of lactoferricin. Mass spectrometry data support the serum stability assay results and allowed us to determine preferred proteolysis sites in the peptides. Furthermore, isothermal titration calorimetry experiments showed that the peptides all had weak interactions with albumin, the most abundant protein in human serum.Taken together, the results provide insight into the behavior of the peptides in human serum and will therefore aid in advancing antimicrobial peptide design towards systemic applications

The Economic Benefits Resulting from the First 8 Years of the Global Programme to Eliminate Lymphatic Filariasis (2000–2007)

Lymphatic filariasis (LF), commonly known as ‘elephantiasis’, is one of the world's most debilitating infectious diseases. In 83 countries worldwide, more than 1.3 billion people are at risk of infection with an estimated 120 million individuals already infected. A recent publication reviewing the health impact of the first 8 years of the Global Programme to Eliminate Lymphatic Filariasis (GPELF) demonstrated the enormous health benefits achieved in populations receiving annual mass drug administration (MDA), as a result of infection prevented, disease progression halted, and ancillary treatment of co-infections. To date, however, no studies have estimated the economic value of these health benefits, either to the individuals or the societies afflicted with LF. Our study estimates that US$21.8 billion will be gained among individuals benefitting from just the first 8 years of the Global Programme, and an additional US$2.2 billion will be saved by the health systems of endemic countries. Treating endemic populations is possible at very low cost – particularly because of the generous drug donations from two pharmaceutical companies – but results in enormous economic benefits. Findings from this study yield a much clearer understanding the GPELF's full economic impact and strengthen the conviction that it is a ‘best buy’ in global health