Using a SMALP platform to determine a sub-nm single particle cryo-EM membrane protein structure

The field of membrane protein structural biology has been revolutionized over the last few years with a number of high profile structures being solved using cryo-EM including Piezo, Ryanodine receptor, TRPV1 and the Glutamate receptor. Further developments in the EM field hold the promise of even greater progress in terms of greater resolution, which for membrane proteins is still typically within the 4-7 angstrom range. One advantage of a cryo-EM approach is the ability to study membrane proteins in more "native" like environments for example proteoliposomes, amphipols and nanodiscs. Recently, styrene maleic acid co-polymers (SMA) have been used to extract membrane proteins surrounded by native lipids (SMALPs) maintaining a more natural environment. We report here the structure of the Escherichia coli multidrug efflux transporter AcrB in a SMALP scaffold to sub-nm resolution, with the resulting map being consistent with high resolution crystal structures and other EM derived maps. However, both the C-terminal helix (TM12) and TM7 are poorly defined in the map. These helices are at the exterior of the helical bundle and form the greater interaction with the native lipids and SMA polymer and may represent a more dynamic region of the protein. This work shows the promise of using an SMA approach for single particle cryo-EM studies to provide sub-nm structures.Peer reviewe

Artificial membranes for membrane protein purification, functionality and structure studies.

Membrane proteins represent one of the most important targets for pharmaceutical companies. Unfortunately, technical limitations have long been a major hindrance in our understanding of the function and structure of such proteins. Recent years have seen the refinement of classical approaches and the emergence of new technologies that have resulted in a significant step forward in the field of membrane protein research. This review summarizes some of the current techniques used for studying membrane proteins, with overall advantages and drawbacks for each method

Membrane Proteins From A to Z Artificial membranes for membrane protein purification, functionality and structure studies

Abstract Membrane proteins represent one of the most important targets for pharmaceutical companies. Unfortunately, technical limitations have long been a major hindrance in our understanding of the function and structure of such proteins. Recent years have seen the refinement of classical approaches and the emergence of new technologies that have resulted in a significant step forward in the field of membrane protein research. This review summarizes some of the current techniques used for studying membrane proteins, with overall advantages and drawbacks for each method

Genética molecular del virus del síndrome reproductor y respiratorio porcino : aspectos evolutivos, diagnósticos e inmunógenos

El síndrome reproductor y respiratorio porcino esta producido por un virus arn, encuadrado en una reciente familia: arteriviridae. Se han desarrollado tres objetivos en este trabajo: 1/ el desarrollo y puesta a punto de una técnica rt-pcr para el diagnostico directo de la enfermedad; 2/ la expresión de la proteína codificada por la orf-5 vírica tanto en sistemas procariotas como eucariotas; 3/ el estudio de variabilidad existente entre cepas europeas del virus. Los resultados del primer objetivo indican que la técnica puesta a punto es altamente sensible y especifica, permitiendo la detección de aproximadamente 100 di50ct en muestras de campo. Con respecto al segundo objetivo, se ha logrado la caracterización de la antes mencionada proteína, así como la purificación parcial de ella. Por ultimo, los estudios de variabilidad demostraron una elevada semejanza entre las cepas analizadas, aunque si bien mayor entre las de un mismo paí