Enzymatically Produced Trimethylamine N-Oxide: Conserving It or Eliminating It
Round 1
Reviewer 1 Report
The manuscript by Catucci, Levi, and co-workers provided a nice review on TMAO, TMA, and FMO. I learned a lot when reading this review article, but sometimes got lost due to the organization of the text. Therefore, I suggest its acception in Catalysts after some minor issues are solved.
The authors need to improve their organization of text. I sometimes cannot find the interconnection between two adjacent paragraphs and do not understand what the authors try to deliver. For instance,
The authors used quite a lot of text (section 2) to describe flavin-containing monooxygenases, even before the detailed introduction of TMA and TMAO. The discussion of TMAU in Section 2.2 (Page 3) and the discussion of unfolded protein response in Section 2.4 (Page 6) are confusing before the readers get a good understanding on the function of TMAO. I would suggest the author to change their paper structure, reviewing both FMO and TMAO first before discussing their relations/connections. I do not see how the discussion of FMO-catalyzed production of taurine (Section 2.4, Page 6) is related to a review article focusing on TMAO.
In addition, most of the quantitative results are presented in text and tables, but not figures. All figures in this review article are illustrations and structures. If there are no copyright concerns, I would suggest the authors to include some key results from other publications, which will definitely enhance the readability of this review article.
Author Response
Comment #1
The authors used quite a lot of text (section 2) to describe flavin-containing monooxygenases, even before the detailed introduction of TMA and TMAO. The discussion of TMAU in Section 2.2 (Page 3) and the discussion of unfolded protein response in Section 2.4 (Page 6) are confusing before the readers get a good understanding on the function of TMAO. I would suggest the author to change their paper structure, reviewing both FMO and TMAO first before discussing their relations/connections. I do not see how the discussion of FMO-catalyzed production of taurine (Section 2.4, Page 6) is related to a review article focusing on TMAO.
Response
We thank the reviewer for the useful comments. We acknowledge that both paragraphs 2.2 and 2.4 needed re-arrangement in the manuscript. The following changes were applied to the manuscript:
1) A short description of the discovery and physiological function of TMAO after scheme 1 to provide a preliminary introduction of the molecule
2) The discussion of TMAU (previously in paragraph 2.2) was moved and enclosed in paragraph 3.2 titled “TMA and its toxicity”. The new location in the manuscript will allow the reader to better understand that TMAU is the accumulation of TMA in the body and that hFMO3 is the enzyme responsible for the detoxification of this compound because the enzymatic reaction is extensively described in the previous section of the manuscript.
3) The discussion of the unfolded protein response (previously in paragraph 2.4) was moved to section 4. This new section, titled “Protein folding: a biochemical process shared by FMO and TMAO” contains two paragraphs:
- 4.1. FMO and protein folding (previously part of “2.4. FMO, endogenous substrates and protein folding homeostasis”)
- 4.2 TMAO as a small chaperone (previously “3.4 Protein folding and TMAO”)
This new section follows the descriptions of FMO (section 2) and TMA/TMAO (section 3). We hope that consequently the reader will have the opportunity to better understand and appreciate the interconnections among the enzyme and the small molecules. The above changes will guide the reader to an overall understanding of the subjects of the review and will lighten the text of paragraph “2.4. Endogenous substrates” (previously “2.4. FMO, endogenous substrates and protein folding homeostasis”). We wish to keep the information about taurine, as we believe that the transformation of the endogenous substrate hypotaurine to taurine by FMO is relevant for this review because it is the most recent discovery on the catalytic activity of the enzyme. As paragraph 2.4 does not include the protein folding homeostasis anymore, the relevance of taurine is even more evident among the other endogenous substrates.
Comment #2
In addition, most of the quantitative results are presented in text and tables, but not figures. All figures in this review article are illustrations and structures. If there are no copyright concerns, I would suggest the authors to include some key results from other publications, which will definitely enhance the readability of this review article.
Response
We believe that for several reasons the quantitative data that are currently mentioned in the text could not be easily illustrated through histograms or other graphical formats. For instance, concentrations of TMAO in body fluids cannot be aggregated in a single figure because they are measured with different methodologies and a statistical aggregation is not feasible in the absence of homogeneous data. Therefore, we would rather keep the data in the current format avoiding the addition of a new figure, that in the case of being from an already published paper could also introduce copyright issues for its reproduction.
Reviewer 2 Report
This review deals with the FMO3 enzyme and its relation to the reactions involves TMA and NADPH. The review starts with classification of the enzyme as respect to structure and function. Then it describes nicely its catalytic activity, and finally its relation to TMA reaction.
The review is well written, I have few comments regarding the figures, and more specifically the figure captions.
Figure 1: define the yellow and green molecules. Maybe add few sentences regarding how the model was created.
Figure 3; Mark yFMO in the figure
Figure 6: which protein was shown? how the models have been done?
Figure 7: what are the different colors of the arrows stand for - define.
Author Response
Comment #1
Figure 1: define the yellow and green molecules. Maybe add few sentences regarding how the model was created.
Response
We have edited the figure legend defining the molecules with the assignment of a color code and we have added a description of how the model was created including appropriated reference.
Comment #2
Figure 3; Mark yFMO in the figure
Response
yFMO now is appropriately labelled in the figure.
Comment #3
Figure 6: which protein was shown? how the models have been done?
Response
The caption of figure 6 now contains the description of the name of the protein that was used with appropriate reference to the .pdb file of the crystal structure. Morever, the caption also includes a reference to the software used for the presentation of the molecules in the figure.
Comment #4
Figure 7: what are the different colors of the arrows stand for - define.
Response
All the colors of the arrows used in the figure are now described in details in the caption.
