PONE-D-24-45122The binding modes of brazilin and hematein from Caesalpinia sappan L.  to Cutibacterium acnes lipase: Simulation studies.PLOS ONE

Dear Dr. Chairatana,

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==============================Please revise the manuscript addressing the concerns of all three reviewers. Reviwer #2 was critical, however, I believe his/her concerns can be addresses during the revision.

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Reviewers' comments:

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Comments to the Author

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

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Reviewer #1: Yes

Reviewer #2: No

Reviewer #3: Yes

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

Reviewer #1: Yes

Reviewer #2: N/A

Reviewer #3: Yes

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

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Reviewer #1: Yes

Reviewer #2: No

Reviewer #3: Yes

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Reviewer #1: No

Reviewer #2: Yes

Reviewer #3: Yes

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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: In reviewing the manuscript, I have noted several instances of spelling errors that should be addressed before publication. These minor issues, while not significantly affecting the overall content, could impact clarity of the manuscript. A careful proofreading of the document is recommended to correct these errors.

Reviewer #2: There are several technical queries with the manuscript mentioned below, which therefore do not allow this manuscript for publishing in its current state.

1. Pg. 13, line 1; default docking protocol; what are those default protocols, it should be mentioned somewhere in the literature (or in the SI).

2. Is “1x1x1 nm3” the size of the box or padding distance, if it’s the size of the box as mentioned, then mention the padding distance?

3. Whether PBC effects were taken into consideration or not?

4. What is the charge of the protein? How many counter ions were added?

5. At what pH was the protonation state of the residues (specially H) decided?

6. How were the ligands parametrized? Which force field, which tool?

7. During energy minimizations was the water relaxed before minimizing the entire system?

8. Since the protocol seems incomplete, can the consistent plateau of minimized energies be provided by the authors in the SI, alongside the equilibrated pressures and temperatures as well?

9. Pg. 15, line 5, the authors say that “This observation is further supported by the electrostatic potential shown in Figure 3C.” but no where in the manuscript is mentioned how the ESPs are generated?

10. Also corroborating the H-bonding observed in dynamics with ESPs does not go well because one is based on force field while the other is quantum mechanical? So, this corroboration must be omitted or must be explained in some other way? Although the authors may take snapshots from the trajectory and perform QM to justify their findings.

11. Why in the figure 4 the superimposition of each ligand at 60ns, 250ns, 410ns, and 500ns, are shown; instead of representing them at consistent intervals. It is like choosing to represent those poses which align well during the dynamics and omitting the rest.

Reviewer #3: Report of Manuscript Number: PONE-D-24-45122

In this study, the authors use docking and molecular dynamics (MD) simulations to investigate the potential application of brazilin and hematein as anti-acne agents. They compare their results with established anti-acne agents, such as KET and TET. Regarding methodology, these methods are commonly used to study such problems. However, there are a couple of concerns regarding the system preparation and the MD methods, which are crucial. Since the work is quite interesting and the results align with the goals of the project, I suggest that the manuscript may be suitable for publication after careful consideration of the following issues.

1) In the abstract, the authors suddenly use abbreviations such as ‘C. sappan L.’ and ‘C. acnes’ without defining them beforehand. It is always better to define abbreviations first, as the authors did for other ligands, such as TET.

2) “Each protein-ligand complex was then placed in a cubic simulation box (dimension of 1x1x1 nm3) and solvated with TIP3P water molecules” This is quite confusing. If the entire box is only 1 nm3 in size and the authors have placed the enzyme within it, the water box would be too small. It is suggested to use a box size of 1 nm3 starting from the protein boundary.

3) What does 0.15 NaCl mean? Are the authors referring to the concentration?

4) I am quite surprised that the authors only performed 1,000 steps for energy minimization. Typically, at least 5,000 steps are recommended. I would also like to see the minimization energy curve to confirm that the system reached the minimum within those 1,000 steps. Moreover, while steepest descent is a powerful algorithm, it is most effective when the system is far from the minimum. As the system approaches the minimum, steepest descent becomes less effective at properly minimizing the system. Therefore, a conjugate gradient algorithm is always recommended after steepest descent to ensure the system reaches a sufficiently localized minimum structure.

5) If the in-house code has already been published, please cite the relevant reference; otherwise, refer to the SI.

6) “This variation in flexibility contributes to the different dynamics and ligand-binding affinities between the two chains.” This statement appears incomplete. Does the author understand the origin of the high flexibility in these new ligands? This is particularly important because the authors’ goal is to conduct a comparative study with an already established ligands.

7) “It is evident that KET and TET undergo significant displacement within the pocket, particularly KET, while the smaller ligands, BRA and HEM, exhibit less mobility.” This is particularly counterintuitive. If a large ligand shows significant displacement within the active site, it implies that the active site is large enough to accommodate the ligand in various orientations. However, this does not necessarily mean that a smaller ligand will be less mobile. Intuitively, the smaller ligand should be even more mobile.

8) “This study serves as a pioneer effort in elucidating the promising capability of HEM to inhibit C. acne lipase.” I would suggest toning down the phrasing a little bit.

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Reviewer #1: Yes:  Vandana Kardam

Reviewer #2: No

Reviewer #3: No

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The binding modes of brazilin and hematein from Caesalpinia sappan L. to Cutibacterium acnes lipase: Simulation studies.

PONE-D-24-45122R1

Dear Dr. Chairatana,

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Kind regards,

Kshatresh Dutta Dubey

Academic Editor

PLOS ONE

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