Role of N-Terminal Extensional Long α-Helix in the Arylesterase from Lacticaseibacillus rhamnosus GG on Catalysis and Stability

Round 1

Reviewer 1 Report

Li et al. reported work "Role of N-Terminal Extensional Long α-Helix in the Arylesterase from Lacticaseibacillus rhamnosus GG on Catalysis and Stability" is well written and can be acceptable for publication in Catalysts upon addressing the below comments. 

Major comment 

1)The author needs to present an inhibitor assay, and a comparison needs to be done with a wild type of Arylestarase activity. Based on these results author can confidently postulate their hypothesis.  

2) Plausible mechanism catalytic activities concerning mutation need to be schematically present in the results and discussion section (i.e., before the conclusion), especially by signifying the importance of N-Terminal Extensional Long α-Helix. 

Minor comments. 

1) Some of the recent references related to mutation-dependent Ayrlestarase catalytic activities need to be cited in the introduction.

2) There are a few typographical mistakes in the manuscript. 

3) Figure 3, 4, and many more need to be specified with the figure legends within the figure. 

4) All the figures were presented in a statistically significant manner. However, I did not see the SD values in the figure captions !!! 

Author Response

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Author Response File: Author Response.pdf

Reviewer 2 Report

1. Every sentence in the abstract and various places in whole manuscript start with Deletion of N-terminal extensional long α-helix. Its hindering in the flow of the paper, please change it ΔN or something similar.

2. LggEst wild type still retained the activities of 90.3% after incubation for 30 min at 45C, but residual activity of the deleted mutant was only 4.6% after incubation for 30 min at 45C? What is the cause of this reduction? Please discuss elaborately in the paper. The reason because the authors shows that thermal unfolding curves showed that the melting temperature Tm of the deleted mutant was similar to that of wild type.

Author Response

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Author Response File: Author Response.pdf

Reviewer 3 Report

Dear Authors,

The strength of your manuscript is the findings of the role of the N-terminal extensional long α-helix on catalytic and stability performance in the L. rhamnosus. The research methodology of this study is reasonable.  However, the concerns of your study are listed below.

1. Many references cited in the manuscript need to be updated. For a high-ranking journal, it should be at most 10 years old. It needs to be better.

2. All sections: for all scientific names, it should “abbreviate the genera name(s)” as this is the second time they have been used. Recheck your manuscript very carefully.

3. Introduction section, lines 36: please explain in more detail and inform in your manuscript, why the esterase and lipase enzymes widely investigated the sequence-structure-function relationship.

4. Line 88, the legend of figure 1: Need reference for the PyMOL software.

5. Materials and Methods section, line 281: Give the full genus name of Escherichia. The abbreviation in line 282 is OK.

6. Materials and Methods section, line 287: It might be better to inform the full name of IPTG and GdnHCl with their abbreviated names.

7. Materials and Methods section, line 297: please explain in brief detail protein purification processes.

8. Materials and Methods section, line 319: please inform all buffer names (pH 4.0-11.0).

9. Materials and Methods section, line 344: use only the abbreviated name.

 

10. Materials and Methods section, line 367: please explain in a brief detail of methods. 

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

The revised manuscript can be accepted for the publication. 

Reviewer 3 Report

After I reviewed this manuscript, I was satisfied with the revised version. Every reviewer's comment was responded to with clear answers.