Chemical crosslinking and mass spectrometry to elucidate the topology of integral membrane proteins
- Debelyy, Mykhaylo O. Division of Biochemistry, Department of Biology, University of Fribourg, Switzerland
- Waridel, Patrice Protein Analysis Facility, Center of Integrative Genomics, Faculty of Biology and Medicine, University of Lausanne, Switzerland
- Quadroni, Manfredo Protein Analysis Facility, Center of Integrative Genomics, Faculty of Biology and Medicine, University of Lausanne, Switzerland
- Schneiter, Roger Division of Biochemistry, Department of Biology, University of Fribourg, Switzerland
- Conzelmann, Andreas Division of Biochemistry, Department of Biology, University of Fribourg, Switzerland
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26.10.2017
Published in:
- PLOS ONE. - 2017, vol. 12, no. 10, p. e0186840
English
Here we made an attempt to obtain partial structural information on the topology of multispan integral membrane proteins of yeast by isolating organellar membranes, removing peripheral membrane proteins at pH 11.5 and introducing chemical crosslinks between vicinal amino acids either using homo- or hetero-bifunctional crosslinkers. Proteins were digested with specific proteases and the products analysed by mass spectrometry. Dedicated software tools were used together with filtering steps optimized to remove false positive crosslinks. In proteins of known structure, crosslinks were found only between loops residing on the same side of the membrane. As may be expected, crosslinks were mainly found in very abundant proteins. Our approach seems to hold to promise to yield low resolution topological information for naturally very abundant or strongly overexpressed proteins with relatively little effort. Here, we report novel XL-MS-based topology data for 17 integral membrane proteins (Akr1p, Fks1p, Gas1p, Ggc1p, Gpt2p, Ifa38p, Ist2p, Lag1p, Pet9p, Pma1p, Por1p, Sct1p, Sec61p, Slc1p, Spf1p, Vph1p, Ybt1p).
- Faculty
- Faculté des sciences et de médecine
- Department
- Département de Biologie
- Language
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- English
- Classification
- Biological sciences
- License
- License undefined
- Identifiers
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- RERO DOC 306171
- DOI 10.1371/journal.pone.0186840
- Persistent URL
- https://folia.unifr.ch/unifr/documents/306238
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