Characterizing glucokinase variant mechanisms using a multiplexed abundance assay
Abstract Background Amino acid substitutions can perturb protein activity in multiple ways. Understanding their mechanistic basis may pinpoint how residues contribute to protein function. Here, we characterize the mechanisms underlying variant effects in human glucokinase (GCK) variants, building on...
| Main Authors: | , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
BMC
2024-04-01
|
| Series: | Genome Biology |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s13059-024-03238-2 |
| _version_ | 1797199416327667712 |
|---|---|
| author | Sarah Gersing Thea K. Schulze Matteo Cagiada Amelie Stein Frederick P. Roth Kresten Lindorff-Larsen Rasmus Hartmann-Petersen |
| author_facet | Sarah Gersing Thea K. Schulze Matteo Cagiada Amelie Stein Frederick P. Roth Kresten Lindorff-Larsen Rasmus Hartmann-Petersen |
| author_sort | Sarah Gersing |
| collection | DOAJ |
| description | Abstract Background Amino acid substitutions can perturb protein activity in multiple ways. Understanding their mechanistic basis may pinpoint how residues contribute to protein function. Here, we characterize the mechanisms underlying variant effects in human glucokinase (GCK) variants, building on our previous comprehensive study on GCK variant activity. Results Using a yeast growth-based assay, we score the abundance of 95% of GCK missense and nonsense variants. When combining the abundance scores with our previously determined activity scores, we find that 43% of hypoactive variants also decrease cellular protein abundance. The low-abundance variants are enriched in the large domain, while residues in the small domain are tolerant to mutations with respect to abundance. Instead, many variants in the small domain perturb GCK conformational dynamics which are essential for appropriate activity. Conclusions In this study, we identify residues important for GCK metabolic stability and conformational dynamics. These residues could be targeted to modulate GCK activity, and thereby affect glucose homeostasis. |
| first_indexed | 2024-04-24T07:15:24Z |
| format | Article |
| id | doaj.art-ae336200514940dba9340a11b8a1aaa3 |
| institution | Directory Open Access Journal |
| issn | 1474-760X |
| language | English |
| last_indexed | 2024-04-24T07:15:24Z |
| publishDate | 2024-04-01 |
| publisher | BMC |
| record_format | Article |
| series | Genome Biology |
| spelling | doaj.art-ae336200514940dba9340a11b8a1aaa32024-04-21T11:20:42ZengBMCGenome Biology1474-760X2024-04-0125112210.1186/s13059-024-03238-2Characterizing glucokinase variant mechanisms using a multiplexed abundance assaySarah Gersing0Thea K. Schulze1Matteo Cagiada2Amelie Stein3Frederick P. Roth4Kresten Lindorff-Larsen5Rasmus Hartmann-Petersen6The Linderstrøm-Lang Centre for Protein Science, Department of Biology, University of CopenhagenThe Linderstrøm-Lang Centre for Protein Science, Department of Biology, University of CopenhagenThe Linderstrøm-Lang Centre for Protein Science, Department of Biology, University of CopenhagenThe Linderstrøm-Lang Centre for Protein Science, Department of Biology, University of CopenhagenDonnelly Centre, University of TorontoThe Linderstrøm-Lang Centre for Protein Science, Department of Biology, University of CopenhagenThe Linderstrøm-Lang Centre for Protein Science, Department of Biology, University of CopenhagenAbstract Background Amino acid substitutions can perturb protein activity in multiple ways. Understanding their mechanistic basis may pinpoint how residues contribute to protein function. Here, we characterize the mechanisms underlying variant effects in human glucokinase (GCK) variants, building on our previous comprehensive study on GCK variant activity. Results Using a yeast growth-based assay, we score the abundance of 95% of GCK missense and nonsense variants. When combining the abundance scores with our previously determined activity scores, we find that 43% of hypoactive variants also decrease cellular protein abundance. The low-abundance variants are enriched in the large domain, while residues in the small domain are tolerant to mutations with respect to abundance. Instead, many variants in the small domain perturb GCK conformational dynamics which are essential for appropriate activity. Conclusions In this study, we identify residues important for GCK metabolic stability and conformational dynamics. These residues could be targeted to modulate GCK activity, and thereby affect glucose homeostasis.https://doi.org/10.1186/s13059-024-03238-2MAVEDMSProtein stabilityProtein dynamicsGCK |
| spellingShingle | Sarah Gersing Thea K. Schulze Matteo Cagiada Amelie Stein Frederick P. Roth Kresten Lindorff-Larsen Rasmus Hartmann-Petersen Characterizing glucokinase variant mechanisms using a multiplexed abundance assay Genome Biology MAVE DMS Protein stability Protein dynamics GCK |
| title | Characterizing glucokinase variant mechanisms using a multiplexed abundance assay |
| title_full | Characterizing glucokinase variant mechanisms using a multiplexed abundance assay |
| title_fullStr | Characterizing glucokinase variant mechanisms using a multiplexed abundance assay |
| title_full_unstemmed | Characterizing glucokinase variant mechanisms using a multiplexed abundance assay |
| title_short | Characterizing glucokinase variant mechanisms using a multiplexed abundance assay |
| title_sort | characterizing glucokinase variant mechanisms using a multiplexed abundance assay |
| topic | MAVE DMS Protein stability Protein dynamics GCK |
| url | https://doi.org/10.1186/s13059-024-03238-2 |
| work_keys_str_mv | AT sarahgersing characterizingglucokinasevariantmechanismsusingamultiplexedabundanceassay AT theakschulze characterizingglucokinasevariantmechanismsusingamultiplexedabundanceassay AT matteocagiada characterizingglucokinasevariantmechanismsusingamultiplexedabundanceassay AT ameliestein characterizingglucokinasevariantmechanismsusingamultiplexedabundanceassay AT frederickproth characterizingglucokinasevariantmechanismsusingamultiplexedabundanceassay AT krestenlindorfflarsen characterizingglucokinasevariantmechanismsusingamultiplexedabundanceassay AT rasmushartmannpetersen characterizingglucokinasevariantmechanismsusingamultiplexedabundanceassay |