Ou X, Blount P, Hoffman R J, Kung C
Laboratory of Molecular Biology and Department of Genetics, University of Wisconsin, 1525 Linden Drive, Madison, WI 53706, USA.
Proc Natl Acad Sci U S A. 1998 Sep 15;95(19):11471-5. doi: 10.1073/pnas.95.19.11471.
MscL is a mechanosensitive channel in bacteria that responds directly to membrane tension by opening a large conductance pore. To determine functionally important residues within this molecule, we have randomly mutagenized mscL, expressed the genes in living bacteria, and screened for gain-of-function mutants with hampered growth. Expression of these genes caused leakage of cytoplasmic solutes on little or no hypo-osmotic stress. In excised patches, the mutant channels gated at membrane tensions that are less than that required for the gating of the wild-type MscL. Hence, the data suggest that the slowed or no-growth phenotype is caused by solute loss because of inappropriate gating of the channel. Most of the mutations mapped to the first transmembrane domain. When this domain is modeled as an alpha-helix, the most severe mutations are substitutions of smaller amino acids (three glycines and one valine) on one facet, suggesting an important role for this structure in MS channel gating.
MscL是细菌中的一种机械敏感通道，通过打开一个大电导孔直接响应膜张力。为了确定该分子内功能重要的残基，我们对mscL进行了随机诱变，在活细菌中表达这些基因，并筛选生长受阻的功能获得型突变体。这些基因的表达在很少或没有低渗应激的情况下导致细胞质溶质泄漏。在切除的膜片中，突变通道在低于野生型MscL门控所需的膜张力下门控。因此，数据表明生长缓慢或无生长表型是由于通道门控不当导致溶质损失所致。大多数突变定位于第一个跨膜结构域。当将该结构域建模为α螺旋时，最严重的突变是一侧较小氨基酸（三个甘氨酸和一个缬氨酸）的取代，表明该结构在MS通道门控中起重要作用。
