The Knotted Protein UCH-L1 Exhibits Partially Unfolded Forms under Native Conditions that Share Common Structural Features with Its Kinetic Folding Intermediates.

The human ubiquitin C-terminal hydrolase, UCH-L1, is an abundant neuronal deubiquitinase that is associated with Parkinson's disease. It contains a complex Gordian knot topology formed by the polypeptide chain alone. Using a combination of fluorescence-based kinetic measurements, we show that UCH-L1 has two distinct kinetic folding intermediates that are transiently populated on parallel pathways between the denatured and native states. NMR hydrogen-deuterium exchange (HDX) experiments indicate the presence of partially unfolded forms (PUFs) of UCH-L1 under native conditions. HDX measurements as a function of urea concentration were used to establish the structure of the PUFs and pulse-labelled HDX NMR was used to show that the PUFs and the folding intermediates are likely the same species. In both cases, a similar stable core encompassing most of the central β-sheet is highly structured and α-helix 3, which is partially formed, packs against it. In contrast to the stable β-sheet core, the peripheral α-helices display significant local fluctuations leading to rapid exchange. The results also suggest that the main difference between the two kinetic intermediates is structure and packing of α-helices 3 and 7 and the degree of structure in β-strand 5. Together, the fluorescence and NMR results establish that UCH-L1 neither folds through a continuum of pathways nor by a single discrete pathway. Its folding is complex, the β-sheet core forms early and is present in both intermediate states, and the rate-limiting step which is likely to involve the threading of the chain to form the 52-knot occurs late on the folding pathway.This work was supported by the National Science Council (99-2911-I-007-034 and 104-2113-M-001-016), National Tsing Hua University and Academia Sinica, Taiwan. S.-T.D.H. was supported by a Career Development Award (CDA- 00025/2010-C) from the International Human Frontier Science Program. The NMR spectra were obtained at the NMR facility of the Department of Chemistry, University of Cambridge and at the Core Facility for Protein Structural Analysis, supported by the National Core Facility Program for Biotechnology, Taiwan.This is the author accepted manuscript. The final version is available from Elsevier via http://dx.doi.org/10.1016/j.jmb.2016.04.00

Women with endometriosis have higher comorbidities: Analysis of domestic data in Taiwan

AbstractEndometriosis, defined by the presence of viable extrauterine endometrial glands and stroma, can grow or bleed cyclically, and possesses characteristics including a destructive, invasive, and metastatic nature. Since endometriosis may result in pelvic inflammation, adhesion, chronic pain, and infertility, and can progress to biologically malignant tumors, it is a long-term major health issue in women of reproductive age. In this review, we analyze the Taiwan domestic research addressing associations between endometriosis and other diseases. Concerning malignant tumors, we identified four studies on the links between endometriosis and ovarian cancer, one on breast cancer, two on endometrial cancer, one on colorectal cancer, and one on other malignancies, as well as one on associations between endometriosis and irritable bowel syndrome, one on links with migraine headache, three on links with pelvic inflammatory diseases, four on links with infertility, four on links with obesity, four on links with chronic liver disease, four on links with rheumatoid arthritis, four on links with chronic renal disease, five on links with diabetes mellitus, and five on links with cardiovascular diseases (hypertension, hyperlipidemia, etc.). The data available to date support that women with endometriosis might be at risk of some chronic illnesses and certain malignancies, although we consider the evidence for some comorbidities to be of low quality, for example, the association between colon cancer and adenomyosis/endometriosis. We still believe that the risk of comorbidity might be higher in women with endometriosis than that we supposed before. More research is needed to determine whether women with endometriosis are really at risk of these comorbidities

Post-translational modifications of voltage-gated sodium channels in chronic pain syndromes.

In the peripheral sensory nervous system the neuronal expression of voltage-gated sodium channels (Navs) is very important for the transmission of nociceptive information since they give rise to the upstroke of the action potential (AP). Navs are composed of nine different isoforms with distinct biophysical properties. Studying the mutations associated with the increase or absence of pain sensitivity in humans, as well as other expression studies, have highlighted Nav1.7, Nav1.8, and Nav1.9 as being the most important contributors to the control of nociceptive neuronal electrogenesis. Modulating their expression and/or function can impact the shape of the AP and consequently modify nociceptive transmission, a process that is observed in persistent pain conditions. Post-translational modification (PTM) of Navs is a well-known process that modifies their expression and function. In chronic pain syndromes, the release of inflammatory molecules into the direct environment of dorsal root ganglia (DRG) sensory neurons leads to an abnormal activation of enzymes that induce Navs PTM. The addition of small molecules, i.e., peptides, phosphoryl groups, ubiquitin moieties and/or carbohydrates, can modify the function of Navs in two different ways: via direct physical interference with Nav gating, or via the control of Nav trafficking. Both mechanisms have a profound impact on neuronal excitability. In this review we will discuss the role of Protein Kinase A, B, and C, Mitogen Activated Protein Kinases and Ca++/Calmodulin-dependent Kinase II in peripheral chronic pain syndromes. We will also discuss more recent findings that the ubiquitination of Nav1.7 by Nedd4-2 and the effect of methylglyoxal on Nav1.8 are also implicated in the development of experimental neuropathic pain. We will address the potential roles of other PTMs in chronic pain and highlight the need for further investigation of PTMs of Navs in order to develop new pharmacological tools to alleviate pain

Effect of Different Manufacturing Methods on the Conflict between Porosity and Mechanical Properties of Spiral and Porous Polyethylene Terephthalate/Sodium Alginate Bone Scaffolds

In order to solve the incompatibility between high porosity and mechanical properties, this study fabricates bone scaffolds by combining braids and sodium alginate (SA) membranes. Polyethylene terephthalate (PET) plied yarns are braided into hollow, porous three dimensional (3D) PET braids, which are then immersed in SA solution, followed by cross-linking with calcium chloride (CaCl2) and drying, to form PET bone scaffolds. Next, SA membranes are rolled and then inserted into the braids to form the spiral and porous PET/SA bone scaffolds. Samples are finally evaluated for surface observation, porosity, water contact angle, compressive strength, and MTT assay. The test results show that the PET bone scaffolds and PET/SA bone scaffolds both have good hydrophilicity. An increasing number of layers and an increasing CaCl2 concentration cause the messy, loose surface structure to become neat and compact, which, in turn, decreases the porosity and increases the compressive strength. The MTT assay results show that the cell viability of differing SA membranes is beyond 100%, indicating that the PET/SA bone scaffolds containing SA membranes are biocompatible for cell attachment and proliferation