Directed evolution methods for overcoming trade‐offs between protein activity and stability

Engineered proteins are being widely developed and employed in applications ranging from enzyme catalysts to therapeutic antibodies. Directed evolution, an iterative experimental process composed of mutagenesis and library screening, is a powerful technique for enhancing existing protein activities and generating entirely new ones not observed in nature. However, the process of accumulating mutations for enhanced protein activity requires chemical and structural changes that are often destabilizing, and low protein stability is a significant barrier to achieving large enhancements in activity during multiple rounds of directed evolution. Here we highlight advances in understanding the origins of protein activity/stability trade‐offs for two important classes of proteins (enzymes and antibodies) as well as innovative experimental and computational methods for overcoming such trade‐offs. These advances hold great potential for improving the generation of highly active and stable proteins that are needed to address key challenges related to human health, energy and the environment.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/154495/1/aic16814_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/154495/2/aic16814.pd

No. 3, A Survey of Historic Pottery Making in Tennessee

https://digitalcommons.memphis.edu/govpubs-tn-dept-environment-conservation-archaeology-research-series/1002/thumbnail.jp

No. 14, Archaeological Investigations at the Carter House State Historic Site, Franklin, Tennessee

https://digitalcommons.memphis.edu/govpubs-tn-dept-environment-conservation-archaeology-investigations/1013/thumbnail.jp

No. 12, An Archaeological Interpretation of the Site of Fort Blount, a 1790s Terrtorial Militia and Federal Military Post, Jackson County, Tennessee

https://digitalcommons.memphis.edu/govpubs-tn-dept-environment-conservation-archaeology-research-series/1011/thumbnail.jp

Proteogenomic Analysis of Bacteria and Archaea: A 46 Organism Case Study

Experimental evidence is increasingly being used to reassess the quality and accuracy of genome annotation. Proteomics data used for this purpose, called proteogenomics, can alleviate many of the problematic areas of genome annotation, e.g. short protein validation and start site assignment. We performed a proteogenomic analysis of 46 genomes spanning eight bacterial and archaeal phyla across the tree of life. These diverse datasets facilitated the development of a robust approach for proteogenomics that is functional across genomes varying in %GC, gene content, proteomic sampling depth, phylogeny, and genome size. In addition to finding evidence for 682 novel proteins, 1336 new start sites, and numerous dubious genes, we discovered sites of post-translational maturation in the form of proteolytic cleavage of 1175 signal peptides. The number of novel proteins per genome is highly variable (median 7, mean 15, stdev 20). Moreover, comparison of novel genes with the current genes did not reveal any consistent abnormalities. Thus, we conclude that proteogenomics fulfills a yet to be understood deficiency in gene prediction. With the adoption of new sequencing technologies which have higher error rates than Sanger-based methods and the advances in proteomics, proteogenomics may become even more important in the future

No. 6, Historical Information Concerning the Fort Blount-Williamsburg Site, Jackson County, Tennessee

https://digitalcommons.memphis.edu/govpubs-tn-dept-environment-conservation-archaeology-investigations/1005/thumbnail.jp