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Ubiquitin (Ub) sorting receptors facilitate the targeting of ubiquitinated membrane proteins into multivesicular bodies (MVBs). Ub-binding domains (UBDs) have been described in several endosomal sorting complexes required for transport (ESCRT). Using available structural information, we have investigated the role of the multiple UBDs within ESCRTs during MVB cargo selection. We found a novel UBD within ESCRT-I and show that it contributes to MVB sorting in concert with the known UBDs within the ESCRT complexes. These experiments reveal an unexpected level of coordination among the ESCRT UBDs, suggesting that they collectively recognize a diverse set of cargo rather than act sequentially at discrete steps.

We report the crystal structure of a bromide-bound form of the D85S mutant of bacteriorhodopsin, bR(D85S), a protein that uses light energy rather than ATP to pump halide ions across the cell membrane. Comparison of the structure of the halide-bound and halide-free states reveals that both displacements of individual side-chain positions and concerted helical movements occur on the extracellular side of the protein. Analysis of these structural changes reveals how this ion pump first facilitates ion uptake deep within the cell membrane and then prevents the backward escape of ions later in the pumping cycle. Together with the information provided by structures of intermediate states in the bacteriorhodopsin photocycle, this study also suggests the overall design principles that are necessary for ion pumping.

The use of hydrated-lipid gels in which the bilayer is an infinitely periodic (or at least continuous), three-dimensional structure offers a relatively new approach for the crystallization of membrane proteins. While excellent crystals of the Halobacterial rhodopsins have been obtained with such media, success remains poor in extending their use to other membrane proteins. Experience with crystallization of bacteriorhodopsin has led us to recognize a number of improvements that can be made in the use of such hydrated-gel media, which may now prove to be of general value for the crystallization of other membrane proteins. © 2003 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 66: 300–316, 2002

A progressive radial neuropathy of unknown etiology despite 1.5T magnetic resonance imaging (MRI) and surgical exploration was identified as an intraneural perineurioma by a localized Tinel's sign, an enlarged radial nerve at the spiral groove by 3.0T MRI, and a fascicular biopsy. The distinction between the initial diagnoses of inflammatory, demyelinating polyneuropathy and perineurioma was made by immunohistochemistry and electron microscopy. A slowly progressing, localized mononeuropathy should include perineurioma in the differential diagnosis.

We present a simple, rapid method for detecting short DNA sequences that combines a novel isothermal amplification method (EXPAR) with visual, colorimetric readout based on aggregation of DNA-functionalized gold nanospheres. The reaction is initiated by a trigger oligonucleotide, synthetic in nature for this proof-of-principle study, which is exponentially amplified at 55 degrees C and converted to a universal reporter oligonucleotide capable of bridging two sets of DNA-functionalized gold nanospheres. This reaction provides10(6)-fold amplification/conversion in under 5 min. When combined with a solution containing DNA nanospheres, the bridging reporter causes nanosphere aggregation. The resulting color change from red to dark purple or blue is enhanced through spotting the solution onto a C18 reversed-phase thin-layer chromatography plate. The reaction can easily be adapted for detection of different trigger oligonucleotides using the same set of DNA nanospheres. It permits detection of as low as 100 fM trigger oligonucleotide in under 10 min total assay time, with minimal reagent consumption and requirement for instrumentation. We expect that combining this simple, versatile assay with trigger generation from a genomic target DNA sequence of interest will be a powerful tool in the development of rapid and simple point-of-care molecular diagnostic applications.

Background and aims: PSC-IBD is reported to represent a distinct phenotype of IBD characterized by colitis, rectal sparing and backwash ileitis, but this has so far not been confirmed in large well-phenotyped cohorts. The aim of this study was to assess the IBD phenotype associated with PSC in a large Dutch PSC cohort using endoscopic and histopathological criteria. Methods: PSC cases were identified and ascertained, fulfilling well-established serological, histological and radiological criteria in 30 hospitals in The Netherlands. IBD location was recorded according to the Montreal classification. To assess the occurrence of backwash ileitis a subgroup analysis was performed in 40 cases and 80 ageand sex-matched IBD controls with at least one complete ileocolonoscopy including terminal ileum histology, reviewing 370 endoscopy and pathology reports written between 2001 and 2010. Results: 324 (64%) of a total of 506 PSC patients had coexistent IBD, mainly ulcerative colitis (UC, 72%). 147 (80%) of the PSC-UC patients had a pancolitis, 28 (15%) a left sided colitis and eight (4%) a proctitis. Sixty (95%) PSC-Crohn's disease (CD) patients had an (ileo)colitis and three ileitis only (5%). In the subgroup analysis of 40 PSC-IBD patients twenty-seven (68%) PSC-UC patients were identified, as well as twelve (30%) PSC-CD patients and one (2%) PSC-undetermined IBD patient. Twenty-five (93%) PSC-UC patients had a pancolitis, compared to thirty-three (61%) matched UC patients (p = 0.034). Left sided colitis was seen in seventeen (32%) UC controls and in none of the PSC-UC patients (p = 0.001). Backwash ileitis was seen in only one (4%) PSC-UC patient and in none of the UC controls. Conclusion: PSC-IBD represents a distinct IBD phenotype. The majority of Dutch PSC-UC patients have a pancolitis. In case of PSC-CD, colonic inflammation is involved in 95% of patients. Backwash ileitis is not a prominent finding in Dutch PSC-UC cases.