TY  - JOUR
A1  - Thess, Andreas
A1  - Hutschenreiter, Silke
A1  - Hofmann, Matthias
A1  - Tampé, Robert
A1  - Baumeister, Wolfgang
A1  - Guckenberger, Reinhard
T1  - Specific orientation and two-dimensional crystallization of the proteasome at metal-chelating lipid interfaces
T2  - Journal of biological chemistry
N2  - The potential of a protein-engineered His tag to immobilize macromolecules in a predictable orientation at metal-chelating lipid interfaces was investigated using recombinant 20 S proteasomes His-tagged in various positions. Electron micrographs demonstrated that the orientation of proteasomes bound to chelating lipid films could be controlled via the location of their His tags: proteasomes His-tagged at their sides displayed exclusively side-on views, while proteasomes His-tagged at their ends displayed exclusively end-on views. The activity of proteasomes immobilized at chelating lipid interfaces was well preserved. In solution, His-tagged proteasomes hydrolyzed casein at rates comparable with wild-type proteasomes, unless the His tags were located in the vicinity of the N termini of α-subunits. The N termini of α-subunits might partly occlude the entrance channel in α-rings through which substrates enter the proteasome for subsequent degradation. A combination of electron micrographs and atomic force microscope topographs revealed a propensity of vertically oriented proteasomes to crystallize in two dimensions on fluid lipid films. The oriented immobilization of His-tagged proteins at biocompatible lipid interfaces will assist structural studies as well as the investigation of biomolecular interaction via a wide variety of surface-sensitive techniques including single-molecule analysis.
Y1  - 2021
UR  - http://publikationen.ub.uni-frankfurt.de/frontdoor/index/index/docId/75998
UR  - https://nbn-resolving.org/urn:nbn:de:hebis:30:3-759984
SN  - 0021-9258
VL  - 277
IS  - 39
SP  - 36321
EP  - 36328
PB  - American Society for Biochemistry and Molecular Biology Publications
CY  - Bethesda, Md
ER  -