Understanding the Inhibition Mechanism of Lignin Adsorption to Cellulase in Terms of Changes in Composition and Conformation of Free Enzymes
, Shuangquan Yao 3 and Hailong Yu 1,3,*
Round 1
Reviewer 1 Report
In this manuscript, the effects of lignin adsorption on the composition and spatial conformation of free enzymes were explored deeply and provided a better understanding of the inhibiting mechanism of lignin adsorption to cellulase. This is a well-written paper containing interesting results which merit publication. There are some problems, which must be solved before it is considered for publication.
1. In Figure 3, an absorption peak appears at 630 cm–1, please indicate to which functional group he belongs.
2. Please cite the necessary references to show that the positions where the four types of C: C1, C2, C3, C4 appear in Figure 4 are accurate.
3. To complete in Table 3, please indicate the pH at the time of zeta potential determination.
Author Response
We thank you for the time, effort, and detailed comments on our manuscript. We have revised the manuscript carefully according to the comments. The point-by-point responses to the comments are listed below.
1. In Figure 3, an absorption peak appears at 630 cm–1, please indicate to which functional group he belongs.
Response: Thanks. The peak at 630 cm-1 was assigned to the bending vibration of the benzene ring. It has been included in the manuscript.
2. Please cite the necessary references to show that the positions where the four types of C: C1, C2, C3, C4appear in Figure 4 are accurate.
Response: Thanks. The reference has been included in the manuscript.
Yang Q, Pan X. Correlation between lignin physicochemical properties and inhibition to enzymatic hydrolysis of cellulose[J]. Biotechnology and bioengineering, 2016, 113: 1213- 1224.
3. To complete in Table 3, please indicate the pH at the time of zeta potential determination.
Response: Thanks. The pH value of 4.8 when measuring zeta potential has been included in the manuscript.
Reviewer 2 Report
Dear Author,
The research paper title "Understanding the inhibition mechanism of lignin adsorption 1 to cellulase in terms of changes in composition and confor- 2 mation of free enzymes" is good in the introduction but needs to correct the methodology and result section in a more scientific way.
Line 33: This line is not clear. Rewrite the sentence
Line 39: The enzyme hydrolysis process need to explain more clearly
Line no 68: Lignin and cellulase both have functional groups,this part should explain with structure
Line no 84: rewrite the sentence
Flow/sequence of contents is not according to the theme, so rewrite
There are lots of grammatical mistakes throughout the paper, check grammatical mistakes and do the correction.
Line no 146:’ The free enzyme solution after 96 h’ not clear so rewrite.
Tables are not complete and unappropriate
Line no 158: Results and discussion portion should rewrite and explain each point
Line no 182: Figure 1. Adsorption curves of cellulase need more clarity
Line no 184: “ endoglu- canase (EG), which randomly cleaves……” not clearly justified , explain again.
Comments for author File: 
Comments.pdf
Author Response
We thank you for the time, effort, and detailed comments on our manuscript. We have revised the manuscript carefully according to the comments. The point-by-point responses to the comments are listed below.
Line 33: This line is not clear. Rewrite the sentence
Response: Thanks. The sentence has been rewritten as “Pretreatment is a process involving physical and/or chemical alterations of lignocellulosic biomass to remove lignin and hemicellulose, making it more suitable for enzymatic hydrolysis.”
Line 39: The enzyme hydrolysis process need to explain more clearly
Response: Thanks. The detailed process of enzymatic hydrolysis had been included in section 3.1.
Line no 68: Lignin and cellulase both have functional groups,this part should explain with structure
Response: Thanks. It has been revised as “Lignin contains carboxyl and sulfonic functional groups, while cellulase contains carboxyl and amino groups. These functional groups can become ionized in water, leading to a solid surface with a net charge. The charges on these functional groups follow the principle of like charges repelling each other and opposite charges attracting each other.”
Line no 84: rewrite the sentence
Flow/sequence of contents is not according to the theme, so rewrite
Response: Thanks. It has been rewritten as “Hydrogen bonding is an important factor in the adsorption of lignin to cellulase. The hydroxyl groups (alcohol and phenolic hydroxyl groups) in lignin can form hydrogen bonds with the free amino or carboxyl groups in cellulase, potentially disrupting the protein's original secondary structure. Hydrogen bonding between the carbonyl and amide groups is responsible for maintaining the stability of the protein's secondary structure.”
There are lots of grammatical mistakes throughout the paper, check grammatical mistakes and do the correction.
Response: Thanks. It has been revised.
Line no 146:’ The free enzyme solution after 96 h’ not clear so rewrite.
Response: Thanks. It has been rewritten.
Tables are not complete and unappropriate
Response: Thanks. It has been revised.
Line no 158: Results and discussion portion should rewrite and explain each point
Response: Thanks. It has been revised.
Line no 182: Figure 1. Adsorption curves of cellulase need more clarity
Response: Thanks. The Figure 1 has been revised.
Line no 184: “ endoglu- canase (EG), which randomly cleaves……” not clearly justified , explain again.
Response: Thanks. It has been revised as “Endoglucanases (EG), can randomly break β-1,4-glycosidic bonds within cellulose molecules, which not only generate two chain ends, but also temporarily loosen the tight binding between these two chain segments and the cellulose molecule.”
Reviewer 3 Report
This manuscript explores the “Understanding the inhibition mechanism of lignin adsorption to cellulase in terms of changes in composition and conformation of free enzymes”. The manuscript is elaborately described and contextualized with the help of previous and present theoretical background. All the references cited are relevant to this area of research. The methods/analytical study are clearly stated. The result and discussion section are clearly presented. The manuscript needs the following modifications before the acceptance.
1. Abstract – Mention your research recommendation.
2. Abstract: Present more results of your work specifically
3. Key words: arrange it in alphabetical order
4. Mention your problem statement clearly.
5. In materials and method section show the materials/experimental photo
6. In Result and discussion section, remove the numbering(1), (2) for sub headings
7. In figures, adjust the font size to manuscript’s font size
8. Conclusion: Mention the research recommendations and scope for the future work
Author Response
We thank you for the time, effort, and detailed comments on our manuscript. We have revised the manuscript carefully according to the comments. The point-by-point responses to the comments are listed below.
1. Abstract – Mention your research recommendation.
Respons: Thanks. It has been included in the manuscript. “It appears that appropriate pretreatment as well as chemical modification of enzymes to resist lignin degradation is a promising long-term pathway to overcome the lignin inhibition during sugar-platform conversion of lignocellulosic bioresources.”
2. Abstract: Present more results of your work specifically
Response: Thanks. The important results has been included in the abstract.
3. Key words: arrange it in alphabetical order
Response: Thanks. It has been revised.
4. Mention your problem statement clearly.
Response: Thanks. It has been revised.
5. In materials and method section show the materials/experimental photo
Response: Thanks. We will show the photos in the future studies.
6. In Result and discussion section, remove the numbering(1), (2) for sub headings
Response: Thanks. It has been revised.
7. In figures, adjust the font size to manuscript’s font size
Response: Thanks. It has been revised.
8. Conclusion: Mention the research recommendations and scope for the future work
Response: Thanks. It has been included in the manuscript.
Reviewer 4 Report
Paper correctly presented with an emphasis on lignin surface chemistry. The usage and interpretation of various physicochemical techniques (FTIR, XPS, CA, etc) was logical and well established.
I recommend acceptance
Author Response
We thank you for the time, effort, and detailed comments on our manuscript.
