Review Reports

Round 1

Reviewer 1 Report

In proposed study authors present application of tagged peptides in protein-protein interaction (PPI) study. Article is well written and highly informative. I recommend to accept the article after minor revision.

1. Binding affinity between the peptides should be included. It will improve the importance of the current study.
2. Correlation study should be performed with microarray, or LC-MS/MS to exhibit the importance of present study.
3. Add following citations in current study to improve the article:  https://doi.org/10.3390/pathogens8040173, https://doi.org/10.3390/biom10010142, https://doi.org/10.1038/nprot.2009.23

Author Response

Response to reviewers

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Reviewer: 1

Comments and Suggestions for Authors

In proposed study authors present application of tagged peptides in protein-protein interaction (PPI) study. Article is well written and highly informative. I recommend to accept the article after minor revision.

Reply: The authors thank very much the reviewer for his time to review the manuscript and his advises to improve it. I carefully followed these advises and I believe that the manuscript is now fairly improved.

1. Binding affinity between the peptides should be included. It will improve the importance of the current study.

Reply: Data on the substrate specificity of BAP1070 and BAP1108 in comparison with Avitag were added to the text in section 4.3 and Supplementry Figure 1S.

2. A correlation study should be performed with microarray, or LC-MS/MS to exhibit the importance of present study.

Reply: Description of LC-MS/MS experiments for BAP1108 was added to the section 4.3. Extracted ion chromatograms and MS/MS spectra were added to Supplementary Figure 2S.

3. Add following citations in current study to improve the article: https://doi.org/10.3390/pathogens8040173, https://doi.org/10.3390/biom10010142, https://doi.org/10.1038/nprot.2009.23

Reply: These citations were added to the text in Discussion part and new section 4.5 of the manuscript.

Author Response File: Author Response.pdf

Reviewer 2 Report

The authors present the experimental workplan with a good modularity for the generation of a new biotin acceptor peptide BAP1108 based on the widely applied original BAP1070. The discussion provides a well documented presentation of the different applications of BAP 1070 for site-specific biotinylation using the PUB method in the field of protein-protein interaction studies.

Since the new peptide BAP1108 differs by 3 aminoacids from the AviTag and by only one from the BAP1070, it would be interesting to have a comment on the expected performance of this new peptide.

line 138: 80-90 kb seems to be the incorrect size

Author Response

Reviewer: 2

Comments and Suggestions for Authors

The authors present the experimental workplan with a good modularity for the generation of a new biotin acceptor peptide BAP1108 based on the widely applied original BAP1070. The discussion provides a well documented presentation of the different applications of BAP 1070 for site-specific biotinylation using the PUB method in the field of protein-protein interaction studies.

Reply: The authors thank very much the reviewer for his time to review the manuscript and his advises to improve it. I carefully followed these advises and I believe that the manuscript is now fairly improved.

1. Since the new peptide BAP1108 differs by 3 aminoacids from the AviTag and by only one from the BAP1070, it would be interesting to have a comment on the expected performance of this new peptide.

Reply: Discussion of properties of the new peptide BAP1108 were added to the text in the section 4.3 and Supplementry Figures 1S and 2S.

2. line 138: 80-90 kb seems to be the incorrect size

Reply: Polyacrylamide gel electrophoresis was used to separate products resulted from annealing and ligation of four smaller oligonucleotides . We cut out the band with 84 bp, corresponding to BAP1070 fragment.

 

Author Response File: Author Response.pdf