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Intra-helical salt bridge contribution to membrane protein insertion
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Science for Life Laboratory, Sweden.ORCID iD: 0000-0003-0568-8281
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Science for Life Laboratory, Sweden.ORCID iD: 0000-0002-7115-9751
(English)Manuscript (preprint) (Other academic)
Abstract [en]

Salt bridges between negatively (D, E) and positively charged (K, R, H) amino acids play an important role in protein stabilization. This has a more prevalent effect in membrane proteins where polar amino acids are exposed to a very hydrophobic environment. In transmembrane (TM) helices the presence of charged residues can hinder the insertion of the helices into the membrane. This can sometimes be avoided by TM region rearrangements after insertion, but it is also possible that the formation of salt bridges could decrease the cost of membrane integration. However, the presence of intra-helical salt bridges in TM domains and their effect on insertion has not been properly studied yet. In this work, we use an analytical pipeline to study the prevalence of charged pairs of amino acid residues in TM α-helices, which shows that potentially salt-bridge forming pairs are statistically over-represented. We then selected some candidates to experimentally determine the contribution of these electrostatic interactions to the translocon-assisted membrane insertion process. Using both in vitro and in vivo systems, we confirm the presence of intra-helical salt bridges in TM segments during biogenesis and determined that they contribute between 0.5-0.7 kcal/mol to the apparent free energy of membrane insertion (ΔGapp). Our observations suggest that salt bridge interactions can be stabilized during translocon-mediated insertion and thus could be relevant to consider for the future development of membrane protein prediction software.

Keywords [en]
electrostatic interactions, membrane insertion, salt bridges, translocon, transmembrane helix
National Category
Bioinformatics (Computational Biology)
Research subject
Biochemistry towards Bioinformatics
Identifiers
URN: urn:nbn:se:su:diva-191210DOI: 10.1101/2021.02.24.432724OAI: oai:DiVA.org:su-191210DiVA, id: diva2:1536955
Available from: 2021-03-12 Created: 2021-03-12 Last updated: 2022-02-25Bibliographically approved
In thesis
1. Transmembrane Proteins and Protein Structure Prediction: What we can learn from Computational Methods
Open this publication in new window or tab >>Transmembrane Proteins and Protein Structure Prediction: What we can learn from Computational Methods
2021 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

A protein’s 3D-structure is essential to understand how proteins function and interact and how biochemical processes proceed in organic life. Despite the advancement in experimental methods, it remains expensive and time-consuming to determine protein structure experimentally. There have been significant advances in machine learning and computational methods where, in many cases, models of protein structure can be determined to a high level of quality. Using computational methods helps predict protein 3D-structure and is often used complementary to experimental methods to give better insight and understanding of biological processes.

This thesis presents studies focusing on the simplicity and transparency of the 3D-structure pipeline. This is done with a new interactive database with full access to the pipeline’s data and code together with tools to analyse and compare models and structures. 

I present a new module for the last step in this pipeline, the final folding of the protein chain, which both simplifies the current pipeline and uses new input data based on the current research. This module predicts better models than its predecessor and produces models more than a magnitude faster than the current state-of-the-art tools. This module also contains a novel way of both folding and docking dimers in one single step. 

There are many examples of how machine learning models contain biases that originate in biased training data, translating into models that do not generalise well. I present a study where experts collaborate to create a high-quality database of Intrinsically Disordered Proteins. Through manual annotation and quality protocols, high-quality training data has been produced that is well suited for machine learning tasks and protein disorder analysis. In this thesis, I also present computational methods pertaining to transmembrane proteins and how they can increase our insight into membrane protein structure. In one study, we use computational methods together with experimental methods to investigate how differently charged residue pairs that form salt bridges inside the membrane of membrane proteins changes the insertion potential. We show that amino acid pairs that form salt bridges in this setting contribute 0.5-0.7 kcal/mol to membrane insertion’s apparent free energy. This gives new insight and advances in how we calculate insertion and can lead to better membrane protein topology predictors. In the final study, we investigate the CPA/AT-transporter family of transmembrane proteins and create a new integrated topology annotation method and structural classification, resulting in new insight into how this family evolved through time. The entire pipeline is published as an interactive database with complete transparency for both the method and data used. The study shows how this family has evolved by duplicating internal regions and how this has caused a structural symmetry in the family. 

This thesis, therefore, contributes to a more accessible and more transparent path of using computational methods to give a more extensive insight into protein structure prediction and how these structures pertain to biochemical processes.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2021. p. 57
Keywords
protein structure prediction, contact prediction, transmembrane protein, topology prediction, machine learning
National Category
Bioinformatics (Computational Biology)
Research subject
Biochemistry towards Bioinformatics
Identifiers
urn:nbn:se:su:diva-191211 (URN)978-91-7911-456-5 (ISBN)978-91-7911-457-2 (ISBN)
Public defence
2021-04-30, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, and online via Zoom, public link is available at the department website, Stockholm, 10:00 (English)
Opponent
Supervisors
Available from: 2021-04-07 Created: 2021-03-12 Last updated: 2022-02-25Bibliographically approved

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Duart, GerardLamb, JohnElofsson, ArneMingarro, Ismael

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