Protein conducting nanopores.

About 50% of the cellular proteins have to be transported into or across cellular membranes. This transport is an essential step in the protein biosynthesis. In eukaryotic cells secretory proteins are transported into the endoplasmic reticulum before they are transported in vesicles to the plasma membrane. Almost all proteins of the endosymbiotic organelles chloroplasts and mitochondria are synthesized on cytosolic ribosomes and posttranslationally imported. Genetic, biochemical and biophysical approaches led to rather detailed knowledge on the composition of the translocon-complexes which catalyze the membrane transport of the preproteins. Comprehensive concepts on the targeting and membrane transport of polypeptides emerged, however little detail on the molecular nature and mechanisms of the protein translocation channels comprising nanopores has been achieved. In this paper we will highlight recent developments of the diverse protein translocation systems and focus particularly on the common biophysical properties and functions of the protein conducting nanopores. We also provide a first analysis of the interaction between the genuine protein conducting nanopore Tom40(SC) as well as a mutant Tom40(SC) (S(54 --> E) containing an additional negative charge at the channel vestibule and one of its native substrates, CoxIV, a mitochondrial targeting peptide. The polypeptide induced a voltage-dependent increase in the frequency of channel closure of Tom40(SC) corresponding to a voltage-dependent association rate, which was even more pronounced for the Tom40(SC) S54E mutant. The corresponding dwelltime reflecting association/transport of the peptide could be determined with t(off) approximately = 1.1 ms for the wildtype, whereas the mutant Tom40(SC) S54E displayed a biphasic dwelltime distribution (t(off)(-1) approximately = 0.4 ms; t(off)(-2) approximately = 4.6 ms).

Insertion of hydrophobic membrane proteins into the inner mitochondrial membrane--a guided tour.

Only a few mitochondrial proteins are encoded by the organellar genome. The majority of mitochondrial proteins are nuclear encoded and thus have to be transported into the organelle from the cytosol. Within the mitochondrion proteins have to be sorted into one of the four sub-compartments: the outer or inner membranes, the intermembrane space or the matrix. These processes are mediated by complex protein machineries within the different compartments that act alone or in concert with each other. The translocation machinery of the outer membrane is formed by a multi-subunit protein complex (TOM complex), that is built up by signal receptors and the general import pore (GIP). The inner membrane houses two multi-subunit protein complexes that each handles special subsets of mitochondrial proteins on their way to their final destination. According to their primary function these two complexes have been termed the pre-sequence translocase (or TIM23 complex) and the protein insertion complex (or TIM22 complex). The identification of components of these complexes and the analysis of the molecular mechanisms underlying their function are currently an exciting and fast developing field of molecular cell biology.