Research & Publications

Courtney E. Gonzalez, Paul Roberts, and Marc Ostermeier, Fitness Effects of Single Amino Acid Insertions and Deletions in TEM-1 β-Lactamase, Journal of Molecular Biology, Volume 431, Issue 12, 2019, Pages 2320-2330 [Science Direct]

Short insertions and deletions (InDels) are a common type of mutation found in nature and a useful source of variation in protein engineering. InDel events have important consequences in protein evolution, often opening new pathways for adaptation. However, much less is known about the effects of InDels compared to point mutations and amino acid substitutions. In particular, deep mutagenesis studies on the distribution of fitness effects of mutations have focused almost exclusively on amino acid substitutions. Here, we present a near-comprehensive analysis of the fitness effects of single amino acid InDels in TEM-1 β-lactamase. While we found InDels to be largely deleterious, partially overlapping deletion-tolerant and insertion-tolerant regions were observed throughout the protein, especially in unstructured regions and at the end of helices. The signal sequence of TEM-1 tolerated InDels more than the mature protein. Most regions of the protein tolerated insertions more than deletions, but a few regions tolerated deletions more than insertions. We examined the relationship between InDel tolerance and a variety of measures to help understand its origin. These measures included evolutionary variation in β-lactamases, secondary structure identity, tolerance to amino acid substitutions, solvent accessibility, and side-chain weighted contact number. We found secondary structure, weighted contact number, and evolutionary variation in class A beta-lactamases to be the somewhat predictive of InDel fitness effects.

Courtney E. Gonzalez and Marc Ostermeier, Pervasive Pairwise Intragenic Epistasis among Sequential Mutations in TEM-1 β-Lactamase, Journal of Molecular Biology, Volume 431, Issue 10, 2019, Pages 1981-1992 [Science Direct]

Interactions between mutations play a central role in shaping the fitness landscape, but a clear picture of intragenic epistasis has yet to emerge. To further reveal the prevalence and patterns of intragenic epistasis, we present a survey of epistatic interactions between sequential mutations in TEM-1 β-lactamase. We measured the fitness effect of ~12,000 pairs of consecutive amino acid substitutions and used our previous study of the fitness effects of single amino acid substitutions to calculate epistasis for over 8000 mutation pairs. Since sequential mutations are prone to physically interact, we postulated that our study would be surveying specific epistasis instead of nonspecific epistasis. We found widespread negative epistasis, especially in beta-strands, and a high frequency of negative sign epistasis among individually beneficial mutations. Negative epistasis (52%) occurred 7.6 times as frequently as positive epistasis (6.8%). Buried residues experienced more negative epistasis that surface-exposed residues. However, TEM-1 exhibited a couple of hotspots for positive epistasis, most notably L221/ R222 at which many combinations of mutations positively interacted. This study is the first to systematically examine pairwise epistasis throughout an entire protein performing its native function in its native host.

Hedieh Saffari, Jeffrey J. Krstyen, Courtney Gonzalez, Frederic C. Clayton, Kristin M. Leiferman, Gerald J. Gleich, Kathryn A. Peterson, Leonard F. Pease, 99mTechnetium-labeled heparin: A new approach to detection of eosinophilic esophagitis–associated inflammation, Journal of Allergy and Clinical Immunology, Volume 132, Issue 6, 2013, Pages 1446-1448 [Science Direct]