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Conditional lethal mutations separate the M13 procoat and Pf3 coat functions of YidC: different YIDC structural
Minyong Chen1, Kun Xie, Nico Nouwen
1Department of Chemistry and Biochemistry Program, The Ohio State University, Columbus, Ohio 43210, USA.
Abstract:
Conditional lethal YidC mutants have been isolated to decipher the role of YidC in the assembly of Sec-dependent and Sec-independent membrane proteins. We now show that the membrane insertion of the Sec-independent M13 procoat-lep protein is inhibited in a short time in a temperature-sensitive mutant when shifted to the nonpermissive temperature. This provides an additional line of evidence that YidC plays a direct role in the insertion of the Sec-independent M13 procoat protein. However, in the temperature-sensitive mutant, the insertion of the Sec-independent Pf3 phage coat protein and the Sec-dependent leader peptidase were not strongly inhibited at the restricted temperatures. Conversely, using a cold-sensitive YidC strain, we find that the membrane insertion of the Sec-independent Pf3 coat protein is blocked, and the Sec-dependent leader peptidase is inhibited at the nonpermissive temperature, whereas the insertion of the M13 procoat protein is nearly normal. These data show that the YidC function for procoat and its function for Pf3 coat and possibly leader peptidase are genetically separable and suggest that the YidC structural requirements are different for the Sec-independent M13 procoat and Pf3 coat phage proteins that insert by different mechanisms.
Insights
YidC protein
Area of Science:
- Molecular Biology
- Protein Biochemistry
- Membrane Protein Biogenesis
Background:
- YidC is a protein essential for membrane protein assembly.
- Its precise role in both Sec-dependent and Sec-independent pathways remains under investigation.
- Understanding YidC's function is crucial for deciphering cellular membrane protein biogenesis.
Purpose of the Study:
- To investigate the specific roles of YidC in the membrane insertion of Sec-independent proteins.
- To differentiate YidC's functions for various membrane proteins using conditional lethal mutants.
- To explore the genetic separability of YidC functions.
Main Methods:
- Utilized temperature-sensitive and cold-sensitive YidC mutants.
- Assessed the membrane insertion of M13 procoat, Pf3 coat, and leader peptidase proteins at restrictive temperatures.
- Analyzed the impact of YidC mutations on Sec-dependent and Sec-independent protein assembly.
Main Results:
- The YidC mutant inhibited M13 procoat insertion at nonpermissive temperatures, confirming YidC's role.
- Pf3 coat and leader peptidase insertion were less affected in the temperature-sensitive mutant.
- Conversely, Pf3 coat insertion was blocked and leader peptidase inhibited in the cold-sensitive mutant, while M13 procoat insertion remained largely unaffected.
- These findings indicate genetically separable functions of YidC.
Conclusions:
- YidC exhibits distinct functional requirements for the membrane insertion of different Sec-independent proteins like M13 procoat and Pf3 coat.
- The study suggests separable YidC functions for M13 procoat and Pf3 coat/leader peptidase.
- Different YidC structural features are likely involved in the insertion of M13 procoat and Pf3 coat proteins, which utilize distinct insertion mechanisms.