ArticleProceedings of the National Academy of Sciences of the United States of America2012
Critical features for biosynthesis, stability, and functionality of a G protein-coupled receptor uncovered by all-versus-all mutations.
Article in Proceedings of the National Academy of Sciences of the United States of America, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 43 papers.
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Who cites it
43 citing papers in PubMed.
- Reactivating the phototransduction cascade with a mutation agnostic gene therapy preserves vision in rod-cone dystrophies.iScience · 2025Article
- Advances in ligand-specific biosensing for structurally similar molecules.Cell systems · 2023Review
- Protein engineering via sequence-performance mapping.Cell systems · 2023Review
- Mutant libraries reveal negative design shielding proteins from supramolecular self-assembly and relocalization in cells.Proceedings of the National Academy of Sciences of the United States of America · 2022Article
- Saturation Mutagenesis of the Transmembrane Region of HokC inInternational journal of molecular sciences · 2021Article
- Directed evolution for high functional production and stability of a challenging G protein-coupled receptor.Scientific reports · 2021Article
- Engineering of Challenging G Protein-Coupled Receptors for Structure Determination and Biophysical Studies.Molecules (Basel, Switzerland) · 2021Review
- Probing the Conformation States of Neurotensin Receptor 1 Variants by NMR Site-Directed Methyl Labeling.Chembiochem : a European journal of chemical biology · 2021Article
- Bridging non-overlapping reads illuminates high-order epistasis between distal protein sites in a GPCR.Nature communications · 2020Article
- MaveDB: an open-source platform to distribute and interpret data from multiplexed assays of variant effect.Genome biology · 2019Article
- Mutations in sigma 70 transcription factor improves expression of functional eukaryotic membrane proteins in Escherichia coli.Scientific reports · 2019Article
- In vivo assembly and large-scale purification of a GPCR - Gα fusion with Gβγ, and characterization of the active complex.PloS one · 2019Article
- Multiplexed assays of variant effects contribute to a growing genotype-phenotype atlas.Human genetics · 2018Review
- PtdIns(4,5)PNature · 2018Article
- Computational design of thermostabilizing point mutations for G protein-coupled receptors.eLife · 2018Article
- Mapping Interaction Sites on Human Chemokine Receptors by Deep Mutational Scanning.Journal of immunology (Baltimore, Md. : 1950) · 2018Article
- A statistical model for improved membrane protein expression using sequence-derived features.The Journal of biological chemistry · 2018Article
- Improving membrane protein expression by optimizing integration efficiency.The Journal of biological chemistry · 2017Article
- Deep sequencing methods for protein engineering and design.Current opinion in structural biology · 2017Review
- Molecular Determinants of Mutant Phenotypes, Inferred from Saturation Mutagenesis Data.Molecular biology and evolution · 2016Article
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6 authors.
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No grant is acknowledged in the PubMed record.
Abstract
The structural features determining efficient biosynthesis, stability in the membrane and, after solubilization, in detergents are not well understood for integral membrane proteins such as G protein-coupled receptors (GPCRs). Starting from the rat neurotensin receptor 1, a class A GPCR, we generated a separate library comprising all 64 codons for each amino acid position. By combining a previously developed FACS-based selection system for functional expression [Sarkar C, et al. (2009) Proc Natl Acad Sci USA 105:14808-14813] with ultradeep 454 sequencing, we determined the amino acid preference in every position and identified several positions in the natural sequence that restrict functional expression. A strong accumulation of shifts, i.e., a residue preference different from wild type, is detected for helix 1, suggesting a key role in receptor biosynthesis. Furthermore, under selective pressure we observe a shift of the most conserved residues of the N-terminal helices. This unique data set allows us to compare the in vitro evolution of a GPCR to the natural evolution of the GPCR family and to observe how selective pressure shapes the sequence space covered by functional molecules. Under the applied selective pressure, several positions shift away from the wild-type sequence, and these improve the biophysical properties. We discuss possible structural reasons for conserved and shifted residues.
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