Protein targeting to the bacterial cytoplasmic membrane

Abstract

Proteins that perform their activity within the cytoplasmic membrane or outside this cell boundary must be targeted to the translocation site prior to their insertion and/or translocation. In bacteria, several targeting routes are known; the SecB- and the signal recognition particle-dependent pathways are the best characterized. Recently, evidence for the existence of a third major route, the twin-Arg pathway, was gathered. Proteins that use either one of these three different pathways possess special features that enable their specific interaction with the components of the targeting routes. Such targeting information is often contained in an N-terminal extension, the signal sequence, but can also be found within the mature domain of the targeted protein. Once the nascent chain starts to emerge from the ribosome, competition for the protein between different targeting factors begins. After recognition and binding, the targeting factor delivers the protein to the translocation sites at the cytoplasmic membrane. Only by means of a specific interaction between the targeting component and its receptor is the cargo released for further processing and translocation. This mechanism ensures the high-fidelity targeting of premembrane and membrane proteins to the translocation site.

FIG. 1

Domain structure of the signal sequence of precursor proteins. (A) Signal sequences of SRP- or SecB-dependent preproteins have a net positive charge in the N region (indicated by +), a hydrophobic H region, and a C region with the signal peptidase cleavage site (✂) preceded by the motif S_nXS_n, in which S_n stands for an amino acid with a small neutral side chain and X stands for any amino acyl residue. (B) Type II signal sequences L_hXS_nC, in which L_h stands for an amino acid with a large hydrophobic side chain and C stands for cysteine. The cleavage site is located between S_n and C. (C) Signal sequences of precursor proteins that are dependent on the twin-Arg route resemble normal signal sequences but have an extended N region and possess the RRXFXK motif, which straddles the H and C domains (9). For both types of signal sequences, the variation in length of the different regions and of the total signal sequence is indicated.

FIG. 2

The SecA- and preprotein-binding sites of SecB are predicted to face in opposite directions in a β-structural conformation. The mainly hydrophobic residues Phe74, Cys76, Val78, and Gln80 are involved in preprotein binding, while the alternating residues Leu75 and Glu77 are involved in SecA binding.

FIG. 3

Coexisting SRP- and SecB-dependent targeting routes. When a preprotein or integral membrane protein emerges from the ribosome, the signal sequence domain is exposed first. (A) When this domain is highly hydrophobic, it will bind SRP and the nascent preprotein will be targeted to the membrane-bound FtsY at the membrane. At the membrane, SRP is released from the preprotein in a GTP-dependent manner. The preprotein or integral membrane protein is subsequently transferred to the translocase, which consists of the integral membrane proteins SecY, SecE, and SecG and the peripherally bound ATPase SecA, or inserts into the membrane via a different pathway. (B) When the signal sequence escapes SRP binding and the mature domain possesses the right features, SecB will bind the mature domain and targets the preprotein to the membrane-bound SecA. At the membrane, SecB donates the preprotein to SecA. Upon the binding of ATP by SecA, translocation is initiated and SecB is released from the ternary complex. (C) When the preprotein is not recognized by SecB, the signal sequence may target it directly to the membrane-bound SecA.