PONE-D-21-14385

Exploring the cytotoxic mechanisms of Pediocin PA-1 towards HeLa and HT29 cells by comparison to known bacteriocins: Microcin E492, Enterocin Heterodimer and Divercin V41.

PLOS ONE

Dear Dr. Buss,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

Please submit your revised manuscript by Jul 18 2021 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

• A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
• A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
• An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

Vivek Gupta

Academic Editor

PLOS ONE

Journal Requirements:

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

When submitting your revision, we need you to address these additional requirements.

1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at

https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and

https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf

2. We suggest you thoroughly copyedit your manuscript for language usage, spelling, and grammar. If you do not know anyone who can help you do this, you may wish to consider employing a professional scientific editing service.  

Whilst you may use any professional scientific editing service of your choice, PLOS has partnered with both American Journal Experts (AJE) and Editage to provide discounted services to PLOS authors. Both organizations have experience helping authors meet PLOS guidelines and can provide language editing, translation, manuscript formatting, and figure formatting to ensure your manuscript meets our submission guidelines. To take advantage of our partnership with AJE, visit the AJE website (http://learn.aje.com/plos/) for a 15% discount off AJE services. To take advantage of our partnership with Editage, visit the Editage website (www.editage.com) and enter referral code PLOSEDIT for a 15% discount off Editage services.  If the PLOS editorial team finds any language issues in text that either AJE or Editage has edited, the service provider will re-edit the text for free.

Upon resubmission, please provide the following:

• The name of the colleague or the details of the professional service that edited your manuscript
• A copy of your manuscript showing your changes by either highlighting them or using track changes (uploaded as a *supporting information* file)
• A clean copy of the edited manuscript (uploaded as the new *manuscript* file)

3. Please amend either the title on the online submission form (via Edit Submission) or the title in the manuscript so that they are identical.

4. Please upload a copy of Figures 1-8, to which you refer in your text. If the figure is no longer to be included as part of the submission please remove all reference to it within the text.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

**********

2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: N/A

Reviewer #2: N/A

**********

3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

**********

5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The qualities that make this research work more interesting are originality of the topic, technical quality, importance in medical field, overall representation and easy understandability, enough figures in line with the text written in the article.

This research work describes very nicely describes application of computational analysis using sequence alignment and modelling tools to investigate the mechanism of action of known bacteriocins.

The title represents manuscript's contents, the conclusions and interpretations are sound, the references are cited properly, this can be considered as unique research as this will help provide a fundamental understanding of cytotoxic mechanisms of Pediocin PA-1 towards HeLa and HT29 cells using molecular simulation tools.

Review and comments are as under -

I have a below recommendation for revisions to improve the readership of the manuscript –

1) Abstract

Line 33 – “Pediocin PA-1 and Microcin E492 and 3D modelling”, please replace “and” with “whereas”

Line 39- 42, Unfortunately, the COVID-19 pandemic meant that we were unable to carry out experiments in the lab, and the unavailability of important data meant we were unable to make solid conclusions but rather suggestions.

Please revise the statement as below –

Unfortunately, due to the COVID-19 pandemic, we were unable to carry out experiments in the lab, and the unavailability of important data we were unable to provide and validate our solid conclusions, but rather suggestions.

Line 42, Remove – “However”, however and despite mean the same. So, please use either one.

2) Introduction

Line 55, please remove “ ‘ “ after diagnosis

Line 56, replace “looks at” with “investigated”

Line 59, replace “ – (Hyphen)“ with a “ , (comma)”

Line 69, replace “looking into” with “discussing”

Line 71, replace “researched in greater detail” with “thoroughly researched or thoroughly evaluated.”

Line 73, remove “been”

Line 78 – 80, “against HT29 – being of the same class of toxin as Pediocin PA-78 1 we hope to identify what is different between these two bacteriocins, and as such gain greater insight into Pediocin PA-1’s mechanism of action.”

Please replace this statement with below –

“against HT29, despite it belonged to the same class of toxin as Pediocin PA-78 1 we hope to identify the difference between these two bacteriocins, and thereby gain greater insight into Pediocin PA-1’s mechanism of action.”

Line 82, please replace “when it has come” with “pertaining”

Line 83, please replace “the structure have been shown to form a disulphide bridge” with “the structure indicated a disulphide bridge formation”

Line 84, please replace “found” with “reported”

Line 86, please remove “been”

Line 86-87, please replace “permeabilization of the lipid bi-layer whilst also inducing

apoptosis by bio-energetic collapse” with “the lipid bi-layer permeabilization whilst inducing

apoptosis as well by bio-energetic collapse”

Line 88, please replace “been shown” with “demonstrated”

Line 90, please replace “has a” with exhibited”

Line 105, please replace “In this way we were able to give a strong argument for further study, as well”, with “In this way we were able to provide a fundamental understanding at molecular level for further investigation, as well”

3) Methodology –

For all the softwares used in this work such as UniProt, MultiAlin and VMD, please provide the version, name of developer, country of origin, etc.

Line 155, please replace “PBD” with “PDB”

Figure 1 is actually a table, so please convert this to a table and replace in-text citation pertaining to this table.

4) Results Microcin E492 –

Line 193, please replace “to the best of our knowledge, the best model” with “it was the only best possible model”

Line 196, please replace “come fold” with “come together to fold”

5) Results – Enterocin –

Line 227, please replace “research has highlighted that Enterocin A and B potentially have different mechanisms of action” with “research has highlighted potentially different mechanisms of actions for Enterocin A and B”

Line 228, please replace “by” with “upon” and “give” with “provide”

Line 240-241, please replace “there were no disulphide bridges detected” with “no disulphide bridges were detected”

6) Divercin V41 (Did you mean – “Results – Divercin V41”,please be consistent)

Line 275, please replace “could go someway to explain why Divercin V41 does not show” with “could explain up to certain extent, the reason for Divercin 41’s inability to”

Line 281, please replace “on analysis” with “upon investigation”

7) Discussion –

Line 308, please add a “comma” between that and when

Line 309, please add a “comma” between cells and necrosis

Line 315, please replace “-“ with “,”

Line 318, please remove “-“ and add a word “with” between alongside and TLR-2.

Line 323 – 324, please replace “limitations of experimental data available, and the COVID-19 restrictions which prevented us from carrying out lab work we were unable to effectively look at this.” With “COVID-19 restrictions the experimental data were not generated to further validate this hypothesis”

Line 341-344, this whole statement is unclear, please re-phrase it for better understanding.

8) Conclusion –

Please draw a conclusion. I was hoping to see a conclusion with a summary of current computational work. Also, state how this work can be extended to future similar work.

Reviewer #2: 1. Page 4, Line 85: Please provide details about the respective protein receptors here.

2. Page 4, Lines 84-91: These statements need further explanation. A clear reason behind the argument that “it is likely Pediocin PA-1 also has a dual mechanism of cytotoxicity” is missing here. Please elaborate.

3. Page 14, Lines 317-320: Authors are recommended to provide details or evidence on the TLR-4 over-expression in HeLa cells and limited expression in human colon cells.

4. Are there any Bacteriocins available for cancer therapy and how far they are successful?

5. It can be understood that authors were unable to carry out experiments in the lab due to the COVID-19 pandemic. However, how far can we rely on bioinformatic tools to while there is unavailability of important data? Please justify.

6. Conclusion needs be further clarified by the authors.

**********

6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: Yes: Harsh Shaiesh Shah

Reviewer #2: Yes: Vineela Parvathaneni

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

Attachments

Attachment

Submitted filename: Harsh Shah - PONE-D-21-14385.docx

PONE-D-21-14385R1

Exploring the cytotoxic mechanisms of Pediocin PA-1 towards HeLa and HT29 cells by comparison to known bacteriocins: Microcin E492, Enterocin Heterodimer and Divercin V41.

PLOS ONE

Dear Dr. Buss,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

While the authors have significantly amended the manuscript, potentially based on reviewers' comments, I do not see a detailed rebuttal letter addressing the comments. All I see is a generic letter with all edits summarized in a paragraph. Please review the comments from first revision, and provide detailed rebuttal when revising the manuscript.

Please submit your revised manuscript by Sep 18 2021 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

• A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
• A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
• An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

Vivek Gupta

Academic Editor

PLOS ONE

Journal Requirements:

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

Additional Editor Comments (if provided):

While the authors have significantly amended the manuscript, potentially based on reviewers' comments, I do not see a detailed rebuttal letter addressing the comments. All I see is a generic letter with all edits summarized in a paragraph. Please review the comments from first revision, and provide detailed rebuttal when revising the manuscript.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

Exploring the cytotoxic mechanisms of Pediocin PA-1 towards HeLa and HT29 cells by comparison to known bacteriocins: Microcin E492, Enterocin Heterodimer and Divercin V41.

PONE-D-21-14385R2

Dear Dr. Buss,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Vivek Gupta

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments: