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Insertion Properties of Marginally Hydrophobic Helices in the LacY Lactose Permease Transporter
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. (Elofsson)
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. (Cristobal)
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. (Elofsson)ORCID iD: 0000-0002-7115-9751
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Transmembrane helices are generally believed to be recognized individually by the translocon based on theirhydrophobicity, but it has been proposed that they could also be recognized as pairs of helices. The fact thatmost transmembrane helices are individually clearly hydrophobic seems to support separate helix insertion,but there are important exceptions where the helices are only borderline hydrophilic, at least according tosequence-based prediction. Conrming these patterns and characterizing their role for insertion of helices isan important part in deciphering membrane protein insertion and folding. Here, we use a combination ofsequence bioinformatics, simplied physical modeling, and experiments to investigate whether helices in theLacY lactose permease transporter are recognized by the translocon, and if not whether helix-helix interactionsmight stabilize their insertion. From the experimentally determined biological hydrophobicity scale, ve out of thetwelve transmembrane segments of LacY are predicted to have low spontaneous insertion, which is qualitativelyconrmed in a simplied simulation model using an implicit membrane environment as well as experimentallyin vitro. For some pairs a small, but signicant, increase in insertion eciency was seen both in the simulationsand in the in vitro system. However, the overall insertion eciency is only marginally increased when pairsof borderline hydrophobic helices are co-inserted, which suggests that translocon-mediated membrane insertionpredominantly recognizes individual helices. It also seems to imply that stabilization of marginally hydrophobichelices - at least for LacY - is a collective eect in the nal folded membrane protein, rather than caused by favorable interactions and hairpin formation during insertion.

Keyword [en]
Membrane protein, transmembrane helix, hydrophobic, insertion, interactions, Monte Carlo
National Category
Biochemistry and Molecular Biology Bioinformatics and Systems Biology
Research subject
URN: urn:nbn:se:su:diva-42792OAI: diva2:351455
Available from: 2010-09-16 Created: 2010-09-14 Last updated: 2014-11-10Bibliographically approved
In thesis
1. Intra- and intermolecular interactions in proteins: Studies of marginally hydrophobic transmembrane alpha-helices and protein-protein interactions.
Open this publication in new window or tab >>Intra- and intermolecular interactions in proteins: Studies of marginally hydrophobic transmembrane alpha-helices and protein-protein interactions.
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Most of the processes in a living cell are carried out by proteins. Depending on the needs of the cell, different proteins will interact and form the molecular machines demanded for the moment. A subset of proteins called integral membrane proteins are responsible for the interchange of matter and information across the biological membrane, the lipid bilayer enveloping and defining the cell. Most of these proteins are co-translationally integrated into the membrane by the Sec translocation machinery.

This thesis addresses two questions that have emerged during the last decade. The first concerns membrane proteins: a number of α-helices have been observed to span the membrane in the obtained three-dimensional structures even though these helices are predicted not to be hydrophobic enough to be recognized by the translocon for integration. We show for a number of these marginally hydrophobic protein segments that they indeed do not insert well outside of their native context, but that their local sequence context can improve the level of integration mediated by the translocon. We also find that many of these helices are overlapped by more hydrophobic segments. We propose, supported by experimental results, that the latter are initially integrated into the membrane, followed by post-translational structural rearrangements. Finally, we investigate whether the integration of the marginally hydrophobic TMHs of the lactose permease of Escherichia coli is facilitated by the formation of hairpin structures. However our combined efforts of computational simulations and experimental investigations find no evidence for this.

The second question addressed in this thesis is that of the interpretation of the large datasets on which proteins that interact with each other in a cell. We have analyzed the results from several large-scale investigations concerning protein interactions in yeast and draw conclusions regarding the biases, strengths and weaknesses of these datasets and the methods used to obtain them.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholms universitet, 2010. 74 p.
membrane proteins, membrane insertion, marginally hydrophobic helix, hydrophobicity, protein-protein interactions, LacY, GltPh
National Category
Biochemistry and Molecular Biology Biochemistry and Molecular Biology
Research subject
urn:nbn:se:su:diva-42856 (URN)978-91-7447-111-3 (ISBN)
Public defence
2010-10-15, Magnelisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
At the time of the doctoral defense the following publications were not published and had a status as follows: Paper 2: In press; Paper 4 Manuscript.Available from: 2010-09-23 Created: 2010-09-16 Last updated: 2014-11-10Bibliographically approved

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Hedin, Linnea ECristobal, SusanaElofsson, ArneLindahl, Erik
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