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Suppressing mutation-induced protein aggregation in mammalian cells by mutating residues significantly displaced upon the original mutation

Paper ID Volume ID Publish Year Pages File Format Full-Text
3127 151 2014 8 PDF Available
Title
Suppressing mutation-induced protein aggregation in mammalian cells by mutating residues significantly displaced upon the original mutation
Abstract

•Protein aggregation mediated by mutation-induced structural changes is proposed.•Cell-based assays are effective in selecting mutants with reduced aggregation.•Mutation of perturbed residues reduces protein aggregation in mammalian cells.•Strategies described here would be a practical option to reduce protein aggregation.

Mutations introduced to wild-type proteins naturally, or intentionally via protein engineering, often lead to protein aggregation. In particular, protein aggregation within mammalian cells has significant implications in the disease pathology and biologics production; making protein aggregation modulation within mammalian cells a very important engineering topic. Previously, we showed that the semi-rational design approach can be used to reduce the intracellular aggregation of a protein by recovering the conformational stability that was lowered by the mutation. However, this approach has limited utility when no rational design approach to enhance conformational stability is readily available. In order to overcome this limitation, we investigated whether the modification of residues significantly displaced upon the original mutation is an effective way to reduce protein aggregation in mammalian cells. As a model system, human copper, zinc superoxide dismutase mutant containing glycine to alanine mutation at position 93 (SOD1G93A) was used. A panel of mutations was introduced into residues substantially displaced upon the G93A mutation. By using cell-based aggregation assays, we identified several novel variants of SOD1G93A with reduced aggregation propensity within mammalian cells. Our findings successfully demonstrate that the aggregation of a mutant protein can be suppressed by mutating the residues significantly displaced upon the original mutation.

Keywords
Protein aggregation; Mammalian cells; Mutation; Protein engineering; Superoxide dismutaseALS, Amyotrophic lateral sclerosis; DMEM, Dulbecco's modified Eagle's medium; EGFP, enhanced green fluorescent protein; FBS, fetal bovine serum; GFP, green fluorescen
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Suppressing mutation-induced protein aggregation in mammalian cells by mutating residues significantly displaced upon the original mutation
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Publisher
Database: Elsevier - ScienceDirect
Journal: Biochemical Engineering Journal - Volume 91, 15 October 2014, Pages 196–203
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us