Protein translocation in Escherichia coli is mediated by the translocase that in its minimal form consists of the protein-conducting channel SecYEG, and the motor protein, SecA. SecYEG forms a narrow pore in the membrane that allows passage of unfolded proteins only. Molecular dynamics simulations suggest that the maximal width of the central pore of SecYEG is limited to 16 Å. To access the functional size of the SecYEG pore, the precursor of outer membrane protein A was modified with rigid spherical tetraarylmethane derivatives of different diameters at a unique cysteine residue. SecYEG allowed the unrestricted passage of the precursor of outer membrane protein A conjugates carrying tetraarylmethanes with diameters up to 18 Å, whereas a 29 Å sized molecule blocked the translocation pore. Translocation of the protein-organic molecule hybrids was strictly proton motive force-dependent and occurred at a single pore. With an average diameter of an unfolded polypeptide chain of 4-6 Å, the pore accommodates structures of at least 22-24 Å, which is vastly larger than the predicted maximal width of a single pore by molecular dynamics simulations.secretion | Sec-system
Small-molecule modulators are very promising tools for the exploration and manipulation of biological systems beyond the limits of genetics. The modern molecular biology toolkit provides a variety of methods that aid in exploring and understanding the structure and function of biological molecules. However, these methods have limitations, especially in the range of changes and responses that they can accomplish. In this regard, chemical modification provides a complementary approach. Variations can be introduced to the protein to confer features not achievable with the 20 amino acids that are genetically encoded. This is especially true when a combination of properties, such as reversibility, tunability, target specificity, sensitivity to external stimuli, and control over the timescale of the effect, are desired all at the same time.A particularly interesting property to control at the molecular level is transport across barriers such as biological membranes, as such control could easily lead to applications in, for example, sensing and detection and drug delivery. This effect has been pursued in a number of studies on the construction of functional nanopores with either synthetic molecules or naturally occurring channels, the latter mainly as b-barrel structures.[1] Herein, we describe the rational design
Small-molecule modulators are very promising tools for the exploration and manipulation of biological systems beyond the limits of genetics. The modern molecular biology toolkit provides a variety of methods that aid in exploring and understanding the structure and function of biological molecules. However, these methods have limitations, especially in the range of changes and responses that they can accomplish. In this regard, chemical modification provides a complementary approach. Variations can be introduced to the protein to confer features not achievable with the 20 amino acids that are genetically encoded. This is especially true when a combination of properties, such as reversibility, tunability, target specificity, sensitivity to external stimuli, and control over the timescale of the effect, are desired all at the same time.A particularly interesting property to control at the molecular level is transport across barriers such as biological membranes, as such control could easily lead to applications in, for example, sensing and detection and drug delivery. This effect has been pursued in a number of studies on the construction of functional nanopores with either synthetic molecules or naturally occurring channels, the latter mainly as b-barrel structures.[1] Herein, we describe the rational design
The crystalline photochromism of a diarylethene pyridyl ligand is applied to the modulation of the electronic environment of a high-spin Fe(II) metal ion.
Gated ion channels are excitable nanopores in biological membranes. They sense and respond to different triggers in nature. The sensory characteristics of these channels can be modified by protein engineering tools and the channels can be functionally reconstituted into synthetic lipid bilayer membranes. The combination of the advances in protein engineering with electrical and/or optical signal detection possibilities makes ion channels perfect detection modules for sensory devices. However, their integration into analytical devices is problematic due to the instability of lipid bilayers. Here, we report on developing a stable sensory chip containing a mechanosensitive channel in a Si/SiO(2) chip with a 3 μm pore. Our new fabrication strategy was straightforward. It required only lithography and dry etching for the pore definition and membrane release and reduced the risk of membrane rupture in the fabrication process. A gated ion channel could be inserted, with the retention of its function, into the pores of Si/SiO(2) chips and be detectable at the single channel level upon activation. Excitable ion channels in stable small pores can serve as very sensitive detectors of specific molecules.
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