Molecular function of the dual-start motif in the lambda S holin

Area of Science:

• Molecular Biology
• Virology
• Membrane Biology

Background:

• The lambda phage S gene encodes two lysis proteins, S105 and S107, differing by two N-terminal amino acids.
• These proteins have opposing roles in viral lysis, with S105 as an effector and S107 as an inhibitor of membrane hole formation.
• The dual-start motif of the S gene is crucial for regulating the timing of viral lysis, known as the 'lysis clock'.

Purpose of the Study:

• To elucidate the molecular mechanism of the lysis clock regulated by the dual-start motif of the lambda phage S gene.
• To investigate the role of N-terminal protein translocation to the periplasm in S protein function.
• To understand how S107 inhibits and S105 promotes membrane hole formation during viral lysis.

Main Methods:

• Utilized protein fusions by linking the M13 procoat protein VIII signal sequence to the N-termini of S105 and S107.
• Assessed protein translocation by measuring signal sequence cleavage by leader peptidase.
• Studied the effect of membrane depolarization on signal sequence cleavage and S protein function.

Main Results:

• Functional S-dependent hole formation required N-terminal signal sequence removal, indicating periplasmic translocation is essential.
• Translocation of S107's N-terminus was approximately threefold slower than S105's N-terminus.
• Membrane depolarization triggered rapid signal sequence cleavage and hole formation by the S107 moiety of the fusion protein.

Conclusions:

• Functional assembly of S proteins necessitates N-terminal translocation to the periplasm.
• S107, with reversed N-terminal topology, inhibits hole formation by interacting with S105.
• Membrane depolarization facilitates S107 translocation, enabling functional assembly and viral lysis.