Understanding the Significance of Biochemistry in the Storage, Handling, Purification, and Sampling of Amphiphilic Mycolactone
Mycolactone, the amphiphilic macrolide toxin secreted by <i>Mycobacterium ulcerans</i>, plays a significant role in the pathology and manifestations of Buruli ulcer (BU). Consequently, it follows that the toxin is a suitable target for the development of diagnostics and therapeutics for...
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| Format: | Article |
| Language: | English |
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MDPI AG
2019-04-01
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| Series: | Toxins |
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| Online Access: | https://www.mdpi.com/2072-6651/11/4/202 |
| _version_ | 1818039103149572096 |
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| author | Jessica Z. Kubicek-Sutherland Dung M. Vu Aaron S. Anderson Timothy C. Sanchez Paul J. Converse Ricardo Martí-Arbona Eric L. Nuermberger Basil I. Swanson Harshini Mukundan |
| author_facet | Jessica Z. Kubicek-Sutherland Dung M. Vu Aaron S. Anderson Timothy C. Sanchez Paul J. Converse Ricardo Martí-Arbona Eric L. Nuermberger Basil I. Swanson Harshini Mukundan |
| author_sort | Jessica Z. Kubicek-Sutherland |
| collection | DOAJ |
| description | Mycolactone, the amphiphilic macrolide toxin secreted by <i>Mycobacterium ulcerans</i>, plays a significant role in the pathology and manifestations of Buruli ulcer (BU). Consequently, it follows that the toxin is a suitable target for the development of diagnostics and therapeutics for this disease. Yet, several challenges have deterred such development. For one, the lipophilic nature of the toxin makes it difficult to handle and store and contributes to variability associated with laboratory experimentation and purification yields. In this manuscript, we have attempted to incorporate our understanding of the lipophilicity of mycolactone in order to define the optimal methods for the storage, handling, and purification of this toxin. We present a systematic correlation of variability associated with measurement techniques (thin-layer chromatography (TLC), mass spectrometry (MS), and UV-Vis spectrometry), storage conditions, choice of solvents, as well as the impact of each of these on toxin function as assessed by cellular cytotoxicity. We also compared natural mycolactone extracted from bacterial culture with synthesized toxins in laboratory (solvents, buffers) and physiologically relevant (serum) matrices. Our results point to the greater stability of mycolactone in organic, as well as detergent-containing, solvents, regardless of the container material (plastic, glass, or silanized tubes). They also highlight the presence of toxin in samples that may be undetectable by any one technique, suggesting that each detection approach captures different configurations of the molecule with varying specificity and sensitivity. Most importantly, our results demonstrate for the very first time that amphiphilic mycolactone associates with host lipoproteins in serum, and that this association will likely impact our ability to study, diagnose, and treat Buruli ulcers in patients. |
| first_indexed | 2024-12-10T07:53:18Z |
| format | Article |
| id | doaj.art-005707b03a6c4fb8825d5999de8ef8ac |
| institution | Directory Open Access Journal |
| issn | 2072-6651 |
| language | English |
| last_indexed | 2024-12-10T07:53:18Z |
| publishDate | 2019-04-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Toxins |
| spelling | doaj.art-005707b03a6c4fb8825d5999de8ef8ac2022-12-22T01:56:58ZengMDPI AGToxins2072-66512019-04-0111420210.3390/toxins11040202toxins11040202Understanding the Significance of Biochemistry in the Storage, Handling, Purification, and Sampling of Amphiphilic MycolactoneJessica Z. Kubicek-Sutherland0Dung M. Vu1Aaron S. Anderson2Timothy C. Sanchez3Paul J. Converse4Ricardo Martí-Arbona5Eric L. Nuermberger6Basil I. Swanson7Harshini Mukundan8Chemistry Division, Los Alamos National Laboratory, Los Alamos, NM 87545, USAChemistry Division, Los Alamos National Laboratory, Los Alamos, NM 87545, USAChemistry Division, Los Alamos National Laboratory, Los Alamos, NM 87545, USABioscience Division, Los Alamos National Laboratory, Los Alamos, NM 87545, USADepartment of Medicine, Johns Hopkins University Center for Tuberculosis Research, Baltimore, MD 21218, USABioscience Division, Los Alamos National Laboratory, Los Alamos, NM 87545, USADepartment of Medicine, Johns Hopkins University Center for Tuberculosis Research, Baltimore, MD 21218, USABioscience Division, Los Alamos National Laboratory, Los Alamos, NM 87545, USAChemistry Division, Los Alamos National Laboratory, Los Alamos, NM 87545, USAMycolactone, the amphiphilic macrolide toxin secreted by <i>Mycobacterium ulcerans</i>, plays a significant role in the pathology and manifestations of Buruli ulcer (BU). Consequently, it follows that the toxin is a suitable target for the development of diagnostics and therapeutics for this disease. Yet, several challenges have deterred such development. For one, the lipophilic nature of the toxin makes it difficult to handle and store and contributes to variability associated with laboratory experimentation and purification yields. In this manuscript, we have attempted to incorporate our understanding of the lipophilicity of mycolactone in order to define the optimal methods for the storage, handling, and purification of this toxin. We present a systematic correlation of variability associated with measurement techniques (thin-layer chromatography (TLC), mass spectrometry (MS), and UV-Vis spectrometry), storage conditions, choice of solvents, as well as the impact of each of these on toxin function as assessed by cellular cytotoxicity. We also compared natural mycolactone extracted from bacterial culture with synthesized toxins in laboratory (solvents, buffers) and physiologically relevant (serum) matrices. Our results point to the greater stability of mycolactone in organic, as well as detergent-containing, solvents, regardless of the container material (plastic, glass, or silanized tubes). They also highlight the presence of toxin in samples that may be undetectable by any one technique, suggesting that each detection approach captures different configurations of the molecule with varying specificity and sensitivity. Most importantly, our results demonstrate for the very first time that amphiphilic mycolactone associates with host lipoproteins in serum, and that this association will likely impact our ability to study, diagnose, and treat Buruli ulcers in patients.https://www.mdpi.com/2072-6651/11/4/202mycolactonestorage and handlingamphiphiliclipoproteins |
| spellingShingle | Jessica Z. Kubicek-Sutherland Dung M. Vu Aaron S. Anderson Timothy C. Sanchez Paul J. Converse Ricardo Martí-Arbona Eric L. Nuermberger Basil I. Swanson Harshini Mukundan Understanding the Significance of Biochemistry in the Storage, Handling, Purification, and Sampling of Amphiphilic Mycolactone Toxins mycolactone storage and handling amphiphilic lipoproteins |
| title | Understanding the Significance of Biochemistry in the Storage, Handling, Purification, and Sampling of Amphiphilic Mycolactone |
| title_full | Understanding the Significance of Biochemistry in the Storage, Handling, Purification, and Sampling of Amphiphilic Mycolactone |
| title_fullStr | Understanding the Significance of Biochemistry in the Storage, Handling, Purification, and Sampling of Amphiphilic Mycolactone |
| title_full_unstemmed | Understanding the Significance of Biochemistry in the Storage, Handling, Purification, and Sampling of Amphiphilic Mycolactone |
| title_short | Understanding the Significance of Biochemistry in the Storage, Handling, Purification, and Sampling of Amphiphilic Mycolactone |
| title_sort | understanding the significance of biochemistry in the storage handling purification and sampling of amphiphilic mycolactone |
| topic | mycolactone storage and handling amphiphilic lipoproteins |
| url | https://www.mdpi.com/2072-6651/11/4/202 |
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