ArticleProtein science : a publication of the Protein Society2019
Homolog comparisons further reconcile in vitro and in vivo correlations of protein activities by revealing over-looked physiological factors.
Article in Protein science : a publication of the Protein Society, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 13 papers.
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Who cites it
13 citing papers in PubMed, 18 citations in OpenAlex.
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- Rheostat positions: A new classification of protein positions relevant to pharmacogenomics.Medicinal chemistry research : an international journal for rapid communications on design and mechanisms of action of biologically active agents · 2020Article
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- Homolog comparisons further reconcile in vitro and in vivo correlations of protein activities by revealing over-looked physiological factors.Protein science : a publication of the Protein Society · 2019Article
- Substitutions at rheostat position 52 of LacI have long-range effects on the LacI conformational landscape.Biophysical chemistryArticle
- A clinically relevant polymorphism in the NaThe Journal of biological chemistryArticle
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Authors and funding
5 authors at 1 institution in 1 country.
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Abstract
To bridge biological and biochemical disciplines, the relationship between in vitro protein biochemical function and in vivo activity must be established. Such studies can (a) help determine whether properties measured in simple, dilute solutions extrapolate to the complex in vivo conditions and (b) illuminate cryptic biological factors that are new avenues for study. We have explored the in vivo-in vitro relationship for chimeras built from LacI/GalR transcription regulators. In prior studies of individual chimeras, amino acid changes that altered in vitro DNA binding affinity exhibited correlated changes in in vivo transcription repression. However, discrepancies arose when the two datasets were compared to each other: Although their DNA binding domains were identical and their in vitro binding affinities spanned the same range, their in vivo repression ranges differed by >50-fold. Here, we determined that the presence of endogenous ligand for one chimera further exacerbated the offset, but that different abilities to simultaneously bind and "loop" two DNA operators resolves the discrepancy. Indeed, results suggest that the lac operon can be looped by even weakly interacting repressor dimers. For looping-competent repressors, we measured in vitro binding to the secondary operator. Surprisingly, this was largely insensitive to amino acid changes in the repressor protein, which reflects altered specificity; this supports the emerging view that the locations of specificity determining positions can be unique to each protein homolog. In aggregate, this work illustrates how a comparative approach can enrich understanding of the in vivo-in vitro relationship and suggest unexpected avenues for future study.
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