Expression, purification and preliminary pharmacological characterization of the Plasmodium falciparum membrane-bound pyrophosphatase type 1

Jianing Liu; Keni Vidilaseris; Niklas G. Johansson; Orquidea Ribeiro; Loïc Dreano; Jari Yli-Kauhaluoma; Henri Xhaard; Adrian Goldman

doi:10.1371/journal.pone.0322756

Peer Review History

Original SubmissionMarch 28, 2025
28 Mar 2025 Author Response https://doi.org/10.1371/journal.pone.0322756.r001
15 Apr 2025 Decision Letter - Ravinder Kumar, Editor PONE-D-25-16760Expression, purification and preliminary pharmacological characterization of the Plasmodium falciparum membrane-bound pyrophosphatase type 1PLOS ONE Dear Dr. Liu, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. Please submit your revised manuscript by May 30 2025 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org . When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. 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China, whereas NJ was funded by the Finnish Pharmaceutical Society, Magnus Ehrnrooth Foundation and the Swedish Cultural Foundation in Finland. We thank V. Manole for technical help with SEC-MALS, and N. Sipari (Viikki Metabolomics Unit-Helsinki Institute of Life Science) for mass spectrometry services. The Biocenter Finland DDCB-HiLIFE infrastructure is thanked for organizing resources in the HX laboratory.” We note that you have provided funding information that is currently declared in your Funding Statement. However, funding information should not appear in the Acknowledgments section or other areas of your manuscript. We will only publish funding information present in the Funding Statement section of the online submission form. Please remove any funding-related text from the manuscript and let us know how you would like to update your Funding Statement. 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Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes Reviewer #2: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: I Don't Know Reviewer #2: Yes ****** 3. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes Reviewer #2: Yes ****** 4. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes Reviewer #2: Yes ****** 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: 1. Yes, Hi5 is clearly giving a higher yield. Data Shows expression analysis was done for four days. Did the authors try a plateau or decline in the expression level. To my understanding it would be good practice to optimize the duration of culture to get an optimal yield. 2. Yes, statistical tools is used but I am not sure if it is used rigorously. (I think the claim should be made carefully for total yield of desired protein and solubility percentage. Particularly in the case of membrane-bound protein). DDM+CHS solubilized more than other used detergents: I hope the authors checked for the maximum extraction of produced protein. I believe some of the protein remains on the membrane and it cannot be resolved during SDS PAGE. Which may affect the image J based determination of protein yield. Hopefully authors have considered it. 3. Yes, the data is available. 4. I believe the manuscript is in presentable form and written in standard English. Reviewer #2: Manuscript titled" Expression, purification and preliminary pharmacological characterization of the Plasmodium falciparum membrane-bound pyrophosphatase type 1" by Jianing is a nice piece of work. The work is important and done cleanely. Overall the manuscript looks OK but need minor revision before provisionally accepted for publication. My comments and suggestions to authors are as below. 1) Draft need minor writing editing 2) In table 1, 2 please add information about tested compound or drugs like CAS no, grade type, purity % etc 3) In gel image please use white background and bands as black for better visibility 4) In Fig 1 A scale bar is missing 5) Information about protein purity is missing ****** 6. PLOS authors have the option to publish the peer review history of their article (what does this mean? ). If published, this will include your full peer review and any attached files. 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Revision 1
28 Apr 2025 Author Response Reviewer #1: 1. Yes, Hi5 is clearly giving a higher yield. Data Shows expression analysis was done for four days. Did the authors try a plateau or decline in the expression level. To my understanding it would be good practice to optimize the duration of culture to get an optimal yield. Thank you for your comment. As shown in Figure 1, protein expression plateaued on Day 3 (48 hours post-transfection), with no significant increase observed on Day 4 (72 hours post-transfection). We did not extend the experiment to Day 5, as the cells began to break down and viability declined noticeably (see figure below). This could decrease the quality of the expressed protein. Based on these observations, we selected Day 4 as the optimal harvest time. 2. Yes, statistical tools is used but I am not sure if it is used rigorously. (I think the claim should be made carefully for total yield of desired protein and solubility percentage. Particularly in the case of membrane-bound protein). DDM+CHS solubilized more than other used detergents: I hope the authors checked for the maximum extraction of produced protein. I believe some of the protein remains on the membrane and it cannot be resolved during SDS PAGE. Which may affect the image J based determination of protein yield. Hopefully authors have considered it. We acknowledge the reviewer concern regarding the determination of solubility percentage.Yes, we took this into careful consideration. To isolate the solubilized protein from the membrane as effectively as possible and to maximize protein yield, we performed ultracentrifugation at 100,000 ×g for 1 hour. A similar method was also used by Liu et al., to screen detergent and calculate protein yield (Liu et al., Methods Mol. Biol.2020). This protocol was consistently applied to all detergents screened, ensuring a fair comparison of their solubilization efficiency. Therefore, we believe the reported percentage of solubilized protein is reliable across all detergents tested. 3. Yes, the data is available 4. I believe the manuscript is in presentable form and written in standard English. Reviewer #2: Manuscript titled" Expression, purification and preliminary pharmacological characterization of the Plasmodium falciparum membrane-bound pyrophosphatase type 1" by Jianing is a nice piece of work. The work is important and done cleanely. Overall the manuscript looks OK but need minor revision before provisionally accepted for publication. My comments and suggestions to authors are as below. 1) Draft need minor writing editing Thank you for your input. We have revised the draft based on the reviewers suggestions. 2) In table 1, 2 please add information about tested compound or drugs like CAS no, grade type, purity % etc We have added additional information about the compounds used in Tables 1 and 2 of the manuscript. 3) In gel image please use white background and bands as black for better visibility We apologize for the confusion. We have reversed the backgroud from black to white in the GFP fluoresence SDS PAGE images. 4) In Fig 1 A scale bar is missing Thanks for pointing this out, now we added the scale bar on the Fig 1A. 5) Information about protein purity is missing Thanks for pointing this out. SDS-PAGE analysis showed a single prominent band corresponding to the expected molecular weight of the target protein, with no detectable impurity bands, suggesting a purity level of >95%. We have included the result in the manuscript. Attachments Attachment Submitted filename: Response to Reviewers_JL.docx https://doi.org/10.1371/journal.pone.0322756.r003
2 May 2025 Decision Letter - Ravinder Kumar, Editor Expression, purification and preliminary pharmacological characterization of the Plasmodium falciparum membrane-bound pyrophosphatase type 1 PONE-D-25-16760R1 Dear Dr. Jianing Liu We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements. Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication. An invoice will be generated when your article is formally accepted. Please note, if your institution has a publishing partnership with PLOS and your article meets the relevant criteria, all or part of your publication costs will be covered. Please make sure your user information is up-to-date by logging into Editorial Manager at Editorial Manager® and clicking the ‘Update My Information' link at the top of the page. If you have any questions relating to publication charges, please contact our Author Billing department directly at authorbilling@plos.org. If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org. Kind regards, Ravinder Kumar, PhD Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: https://doi.org/10.1371/journal.pone.0322756.r004
Formally Accepted
Acceptance Letter - Ravinder Kumar, Editor PONE-D-25-16760R1 PLOS ONE Dear Dr. Liu, I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now being handed over to our production team. At this stage, our production department will prepare your paper for publication. This includes ensuring the following: * All references, tables, and figures are properly cited * All relevant supporting information is included in the manuscript submission, * There are no issues that prevent the paper from being properly typeset You will receive further instructions from the production team, including instructions on how to review your proof when it is ready. Please keep in mind that we are working through a large volume of accepted articles, so please give us a few days to review your paper and let you know the next and final steps. Lastly, if your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org. If we can help with anything else, please email us at customercare@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Dr. Ravinder Kumar Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0322756.r005

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