SecA Cotranslationally Interacts with Nascent Substrate Proteins In Vivo

Area of Science:

• Bacterial protein secretion
• Molecular cell biology
• Biochemistry

Background:

• SecA is crucial for bacterial protein transport across the cytoplasmic membrane.
• Protein translocation is often a posttranslational process.
• SecA's interaction with ribosomes and nascent chains is under investigation.

Purpose of the Study:

• To investigate the in vivo interaction of SecA with nascent polypeptides.
• To determine if SecA recognizes Sec substrates during their synthesis.
• To elucidate the temporal order of interactions between SecA, ribosomes, and other factors like SecB and trigger factor.

Main Methods:

• Photo-cross-linking of SecA to ribosomes in vivo.
• Identification and microarray analysis of copurifying mRNAs.
• 2-D gel electrophoresis of radioactively labeled nascent polypeptides associated with SecA.
• In vitro analysis of SecA interaction with purified ribosomes and nascent chains.

Main Results:

• SecA was found to interact with nascent polypeptides in vivo, which are confirmed Sec substrates.
• SecA's interaction with nascent chains is largely independent of SecB and trigger factor.
• SecB's interaction with nascent chains is dependent on SecA-ribosome interaction, suggesting SecA acts first.
• SecA can interact with nascent chains as short as ~110 amino acids, earlier than SecB.

Conclusions:

• SecA cotranslationally recognizes nascent Sec substrates.
• This early recognition facilitates efficient targeting of proteins to the Sec machinery.
• The findings suggest a revised model for the initiation of bacterial protein translocation.