Expression and Aggregation of Recombinant Human Consensus Interferon-α Mutant by Pichia pastoris

A recombinant human consensus interferon-α mutant (cIFN) was expressed in Pichia pastoris. The maximum dry cell weight, cIFN concentration and antiviral activity were 160 g l⁻¹, 1.24 g l⁻¹ and 4.1 × 10⁷ IU ml⁻¹, respec tively. The cIFN secreted into the medium was in the form of aggregates dominantly by non-covalent interaction and partially by disulphide bond. When the fermentation supernatant was disaggregated with 6 M guanidine hydrochloride, the antiviral activity of cIFN achieved 2.2 × 10⁸ IU ml⁻¹.     

向我饲养的动物学习——访热带小猴馆饲养员

当我们和其他生命朝夕相处时,也许会重新思考自己和它们之间的位置与关系。动物园的饲养员就是这样,我们经常会听到他们说：“是动物教育了我。”[编者按]     

GAzer: gene set analyzer

Gene Set Analyzer (GAzer) is a web-based integrated gene set analysis tool covering previously reported parametric and non-parametric models. Based on a simulation test for the reported algorithms, we classified and implemented three main statistical methods consisting of the z-statistic, gene permutation and sample permutation for ten gene set categories including Gene Ontology (GO) for human, mouse, rat and yeast. This tool identifies significantly altered gene sets scored by z-statistics and P-values from the z-test or permutation test and provides q-values and Bonferroni P-values to correct multiple hypothesis testing. GAzer allows users to observe changes in expression of each gene in a gene set or to see the significance of the gene sets containing a gene(s) of interest, thus allowing interactive data analysis both at the gene and gene set level. Moreover, GAzer offers extensive annotation for each gene. AVAILABILITY: The GAzer gene set analyzer is freely available at http://integromics.kobic.re.kr/GAzer/. SUPPLEMENTARY INFORMATION: This can be found on the web page (http://integromics.kobic.re.kr/GAzer/supplement.jsp).     

Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes

Research in autophagy continues to accelerate,(1) and as a result many new scientists are entering the field. Accordingly, it is important to establish a standard set of criteria for monitoring macroautophagy in different organisms. Recent reviews have described the range of assays that have been used for this purpose.(2,3) There are many useful and convenient methods that can be used to monitor macroautophagy in yeast, but relatively few in other model systems, and there is much confusion regarding acceptable methods to measure macroautophagy in higher eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers of autophagosomes versus those that measure flux through the autophagy pathway; thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from fully functional autophagy that includes delivery to, and degradation within, lysosomes (in most higher eukaryotes) or the vacuole (in plants and fungi). Here, we present a set of guidelines for the selection and interpretation of the methods that can be used by investigators who are attempting to examine macroautophagy and related processes, as well as by reviewers who need to provide realistic and reasonable critiques of papers that investigate these processes. This set of guidelines is not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to verify an autophagic response.     

A method to determine whether a musculoskeletal model can resist arbitrary external loadings within a prescribed range

Computational models of the musculoskeletal system are prone to design errors. It is possible to create a model that cannot satisfy equilibrium conditions for a set of external loading conditions. A model is ‘loadable’ if there exists a set of muscle forces that can resist an arbitrary applied force within a prescribed range. In this study, a novel mathematical method is introduced to determine whether models are loadable. In addition, an idealised musculoskeletal model is presented in order to develop the theory behind the mathematical method. The method uses the simplex algorithm to determine feasibility of the linear programming problem and can determine loadability for an arbitrary, continuous range of external forces. The method was applied to a three-dimensional model of the shoulder and correctly determined loadability for a range of externally applied forces.     

Identification of fragments targeting an alternative pocket on HIV-1 gp41 by NMR screening and similarity searching

The HIV-1 envelope glycoprotein gp41 fusion intermediate is a promising drug target for inhibiting viral entry. However, drug development has been impeded by challenges inherent in mediating the underlying protein–protein interaction. Here we report on the identification of fragments that bind to a C-terminal sub-pocket adjacent to the well-known hydrophobic pocket on the NHR coiled coil. Using a specifically designed assay and ligand-based NMR screening of a fragment library, we identified a thioenylaminopyrazole compound with a dissociation constant of ～500 μM. Interaction with the C-terminal sub-pocket was confirmed by paramagnetic relaxation enhancement NMR experiments, which also yielded the binding mode. Shape-based similarity searching detected additional phenylpyrazole and phenyltriazole fragments within the library, enriching the hit rate over random screening, and revealing molecular features required for activity. Discovery of the novel scaffolds and binding mechanism suggests avenues for extending the interaction surface and improving the potency of a hydrophobic pocket binding inhibitor.     

Method for Quantitative Study of Airway Functional Microanatomy Using Micro-Optical Coherence Tomography

We demonstrate the use of a high resolution form of optical coherence tomography, termed micro-OCT (μOCT), for investigating the functional microanatomy of airway epithelia. μOCT captures several key parameters governing the function of the airway surface (airway surface liquid depth, periciliary liquid depth, ciliary function including beat frequency, and mucociliary transport rate) from the same series of images and without exogenous particles or labels, enabling non-invasive study of dynamic phenomena. Additionally, the high resolution of μOCT reveals distinguishable phases of the ciliary stroke pattern and glandular extrusion. Images and functional measurements from primary human bronchial epithelial cell cultures and excised tissue are presented and compared with measurements using existing gold standard methods. Active secretion from mucus glands in tissue, a key parameter of epithelial function, was also observed and quantified.     

The lack of effect of pentoxifylline on random skin flap survival

The effects of pentoxifylline on skin flap survival were studied in rabbits. A total of 40 rabbits had caudally based single-pedicle flaps measuring 4 x 14 cm raised on the mid dorsum of each animal. Twenty of these rabbits were given intraperitoneal injections of pentoxifylline in doses of 24 mg/kg per day beginning 48 hours prior to flap construction and continued daily for 7 days postoperatively. The remaining 20 control rabbits received intraperitoneal injections of saline in equal volumes as the experimental groups. At the end of 7 days, viable flap length was visually inspected and measured in all 40 rabbits. There was no significant difference in skin flap viability in rabbits treated with pentoxifylline compared to the control group.     

Rapid Test for Urease and Phenylalanine Deaminase Production

A rapid urea-phenylalanine medium was effective for the identification of Proteus and, with one exception, Providencia. Most Klebsiella and a few Enterobacter were urease-positive with this method.