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A single mutant (G131R) constructed previously showing increased affinity to both DRs and a double mutant (N199R; K201H), showing a decrease in affinity to DR5 have been combined before (to be published elsewhere), and were therefore incorporated in the new combination variants.
Mutation S159R was incorporated in all the combination variants, showing a high affinity ratio change (Supplementary Table 1).
Two combination variants showing the highest affinity increase to DR4 and affinity decrease for DR5 were selected for further studies.
The characteristics of these single mutants were used for rational design of nine combination variants, containing between two and six mutations each.
Comparison between single variants and combination variants in BJAB cell lines showed that 4C7 displays a greatly improved capability to induce cell death when compared with rhTRAILWT, to the single variants and to a combination of three mutations (Supplementary Figure S3).
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Interestingly, another combination variant (4C6: R149I/showedS215D) showed not only lowered binding to DR5, but also decreased binding affinity to both decoy receptors in solution (Supplementary Figure S2).
Counts are stratified by each combination of two variants; for each combination, variant 1 (v1) is the variant in the gene listed first and (v2) is the variant in the gene listed second; The total counts for LOAD patients and controls are given underneath the stratified counts.
Thus, it is possible that S versus LEW is not the only strain combination with variants of Adamts16 representing candidates for the quantitative trait of BP.
For example, a larger pedigree would afford more power to identify individuals sharing not only the same rare structural variant, but in some cases, the same combination of variants involving different genes or intergenic regions.
Population-based phasing methods are probabilistic, limited to generation of short haplotype blocks, and will incorrectly phase rare combinations of variants, exactly those combinations most likely to be medically important.
In a family, the combination of variants is probably specific to that one family, and because of this clustering of variants it will increase the chance to find it.
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com