ORCID Profile
0000-0001-6662-9301
Current Organisation
University of Tokyo
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Publisher: Elsevier BV
Date: 2016
DOI: 10.1016/J.BBAMEM.2015.10.010
Abstract: The mechanosensitive (MS) channel MscCG from the soil bacterium Corynebacterium glutamicum functions as a major glutamate exporter. MscCG belongs to a subfamily of the bacterial MscS-like channels, which play an important role in osmoregulation. To understand the structural and functional features of MscCG, we investigated the role of the carboxyl-terminal domain, whose relevance for the channel gating has been unknown. The chimeric channel MscS-(C-MscCG), which is a fusion protein between the carboxyl terminal domain of MscCG and the MscS channel, was examined by the patch cl technique. We found that the chimeric channel exhibited MS channel activity in Escherichia coli spheroplasts characterized by a lower activation threshold and slow closing compared to MscS. The chimeric channel MscS-(C-MscCG) was successfully reconstituted into azolectin liposomes and exhibited gating hysteresis in a voltage-dependent manner, especially at high pipette voltages. Moreover, the channel remained open after releasing pipette pressure at membrane potentials physiologically relevant for C. glutamicum. This contribution to the gating hysteresis of the C-terminal domain of MscCG confers to the channel gating properties highly suitable for release of intracellular solutes.
Publisher: Springer Science and Business Media LLC
Date: 27-08-2018
DOI: 10.1038/S41598-018-31219-6
Abstract: MscCG, a mechanosensitive channel of Corynebacterium glutamicum provides a major export mechanism for glutamate in this Gram-positive bacterium, which has for many years been used for industrial production of glutamate and other amino acids. The functional characterization of MscCG is therefore, of great significance to understand its conductive properties for different amino acids. Here we report the first successful giant spheroplast preparation of C. glutamicum amenable to the patch cl technique, which enabled us to investigate mechanosensitive channel activities of MscCG in the native membrane of this bacterium. Single channel recordings from these spheroplasts revealed the presence of three types of mechanosensitive channels, MscCG, MscCG2, and CgMscL, which differ largely from each other in their conductance and mechanosensitivity. MscCG has a relatively small conductance of ~340 pS followed by an intermediate MscCG2 conductance of ~1.0 nS and comparably very large conductance of 3.7 nS exhibited by CgMscL. By applying Laplace’s law, we determined that very moderate membrane tension of ~5.5 mN/m was required for half activation of MscCG compared to ~12 mN/m required for half activation of both MscCG2 and CgMscL. Furthermore, by combining the micropipette aspiration technique with molecular dynamics simulations we measured mechanical properties of the C. glutamicum membrane, whose area elasticity module of K A ≈ 15 mN/m is characteristic of a very soft membrane compared to the three times larger area expansion modulus of K A ≈ 44 mN/m of the more elastic E. coli membrane. Moreover, we demonstrate that the “soft” properties of the C. glutamicum membrane have a significant impact on the MscCG gating characterized by a strong voltage-dependent hysteresis in the membrane of C. glutamicum compared to a complete absence of the hysteresis in the E. coli cell membrane. We thus propose that MscCG has evolved and adapted as an MscS-like channel to the mechanical properties of the C. glutamicum membrane enabling the channel to specialize in transport of amino acids such as glutamate, which are major osmolytes helping the bacterial cells survive extreme osmotic stress.
Publisher: Springer Science and Business Media LLC
Date: 12-09-2018
Publisher: Elsevier BV
Date: 12-2020
No related grants have been discovered for Hisashi Kawasaki.