Using Adsorption Energy Distribution for Parameter Estimation of Competitive Cofactor Coupled Enzyme Reaction

Round 1

Reviewer 1 Report

“Using Adsorption Energy Distribution for Parameter Estimation of Competitive Cofactor Coupled Enzyme Reaction " by Waluga et al described the use of Adsorption Energy Distribution (AED) for determining the individual parameters of the reaction kinetics. It shows that this calculation tool can obtain similar parameters as those from experimentation data via the use of Michalis-Menten equation. The results presented support the conclusion that the use of AED can significantly reduce time and cost for determining those kinetic parameters. However, this reviewer is unable to comment on how this AED did the calculation.

Author Response

We regret that the calculation method is partly unclear. We have attached a scheme of the algorithm as Scheme 1 to the SI. We hope that the calculation method will now be clearer. In addition, we decided to publish further data as gitlab project, with a link in the section “Data Availability Statement”. As the time for the revision is quite short, we will publish this as ongoing (e.g. only focusing on calculation of distribution, but no further automatized analysis) but we will be polishing this gitlab project within the next weeks.

Reviewer 2 Report

The manuscript can be published after a major revision. The authors should go through the following points and revise the manuscript.

1. The authors report that AED is superior to NLR. Please make a table and compare AED with NLR mentioning key advantages and disadvantages of the mentioned approaches.

2.       The ADH used in the study is procured from commercial source. Mention the details about the source, initial activity, cost etc. Such data would be needed for reproducibility pusposes.

3.       s. cerevisiae: the s should be capital.

4.       All experiments were performed as triplets. What is the experimental error in this study?

5.       Did the authors observed enzyme inhibition due to alcohols? Have the authors quantified it and used it in the calculation?

6.       Abstract and conclusions should contain key findings supported by data. Please revise it contain too much text.

Minor editing would be necessary for abstract and conclusion

Author Response

1. Thank you for pointing this out. We have added a list to the manuscript so that the benefits are now clearer.

2. We have added the full product number as well as the batch number. With the numbers now given, all relevant data can be obtained from the Sigmaaldrich homepage.

3. Thanks. We have corrected the error at all positions.

4. Thank you for the note. We have added the missed standard deviation to tables S1 and S2.

5. Currently we do not consider inhibition, as this would extend the AED analysis to another dimension. However, this extension is part of our future research. For now, we are focusing on establishing the AED for enzyme kinetic analysis. A Ki value could be calculated according to equation 7, but 3 data points do not justify a calculation of Ki.

6. We have revised the abstract and the conclusions.

Round 2

Reviewer 1 Report

NA

Reviewer 2 Report

The manuscript can be accepted for publication.

Minor language editing is necessary.