A Work in Progress: The Lived Experiences of Black Male Undergraduates at One Predominantly White University

This exploratory study examines the lived academic and social experiences of current black male undergraduate students, including their perceived barriers and their strategies for persistence and achieving success. Study participants included black male undergraduate students at one predominantly white, four-year, public research university. Data collection methods included two focus group interviews with a purposeful sample of 12 undergraduate, black male students. Students reported that explicit and implicit messages of racial hatred have contributed to a general campus atmosphere of discomfort for black male students. The students described an environment where they continually encounter racial micro aggressions and prejudice. Students reported that persisting at Mid-South has been challenging for them. Nevertheless, they have persisted. Among their lessons learned is that networking is powerful; both inter- and intra-racial networking

Directed evolution of cytochrome c for carbon–silicon bond formation: Bringing silicon to life

Organic compounds containing silicon are important for a number of applications, from polymers to semiconductors. The catalysts used for creating carbon-silicon bonds, however, often require expensive trace metals or have limited lifetimes. Borrowing from the ability of some metallo-enzymes to catalyze other rare carbene insertion reactions, Kan et al. used heme proteins to form carbon-silicon bonds across a range of conditions and substrates (see the Perspective by Klare and Oestreich). Directed evolution experiments using cytochrome c from Rhodothermus marinus improved the reaction to be 15 times more efficient than industrial catalysts

Machine learning-assisted directed protein evolution with combinatorial libraries

To reduce experimental effort associated with directed protein evolution and to explore the sequence space encoded by mutating multiple positions simultaneously, we incorporate machine learning in the directed evolution workflow. Combinatorial sequence space can be quite expensive to sample experimentally, but machine learning models trained on tested variants provide a fast method for testing sequence space computationally. We validate this approach on a large published empirical fitness landscape for human GB1 binding protein, demonstrating that machine learning-guided directed evolution finds variants with higher fitness than those found by other directed evolution approaches. We then provide an example application in evolving an enzyme to produce each of the two possible product enantiomers (stereodivergence) of a new-to-nature carbene Si-H insertion reaction. The approach predicted libraries enriched in functional enzymes and fixed seven mutations in two rounds of evolution to identify variants for selective catalysis with 93% and 79% ee. By greatly increasing throughput with in silico modeling, machine learning enhances the quality and diversity of sequence solutions for a protein engineering problem.Comment: Corrected best S-selective variant sequence in Figure 4. Corrected less R-selective variant sequences from Round II Input library in Table 2 and Supp Table 4. Corrections may also be found on PNAS version https://www.pnas.org/content/early/2019/12/26/192177011

Stereodivergent cyclopropanation of unactivated alkenes with heme proteins

Cyclopropyl motifs are present in a variety of compounds important to pharmaceutical, agrochemical, and fragrance industries. The asymmetric synthesis of cyclopropanes is often performed under harsh conditions with toxic, precious metal chiral catalysts. In 2013, the first example of biocatalytic alkene cyclopropanation was reported, using an engineered cytochrome P450 enzyme [1]. Since then, several heme proteins were reported to cyclopropanate a variety of styrenyl alkenes [2], but none have been shown to asymmetrically cyclopropanate more challenging substrates such as unactivated, aliphatic alkenes using the native iron-heme cofactor. Here we report that heme proteins can cyclopropanate unactivated alkenes and that stereoselectivity and activity can be tuned by directed evolution. A few rounds of site-saturation mutagenesis and screening yielded four protein variants with high enantio- and diastereoselectivity for complementary isomers, enabling stereodivergent synthesis of aliphatic cyclopropanes. These iron-porphyrin proteins are fully genetically encoded, and the reactions can be performed under mild, aqueous conditions with whole cells or purified protein. The protein enhances the activity of the native iron-heme cofactor, giving access to a broad array of cyclopropanated products. This example showcases the ability to quickly and efficiently engineer proteins for non-natural biocatalytic function. [1] P.S. Coelho, E.M. Brustad, A. Kannan, F.H. Arnold, Olefin cyclopropanation via carbene transfer catalyzed by engineered cytochrome P450 enzymes., Science. 339 (2013) 307–10. [2] O.F. Brandenberg, R. Fasan, F.H. Arnold, Exploiting and engineering hemoproteins for abiological carbene and nitrene transfer reactions, Curr. Opin. Biotechnol. 38 (2017) in press

Impact of cyclooxygenase inhibitors in the Women's Health Initiative hormone trials: secondary analysis of a randomized trial.

OBJECTIVES: We evaluated the hypothesis that cyclooxygenase (COX) inhibitor use might have counteracted a beneficial effect of postmenopausal hormone therapy, and account for the absence of cardioprotection in the Women's Health Initiative hormone trials. Estrogen increases COX expression, and inhibitors of COX such as nonsteroidal anti-inflammatory agents appear to increase coronary risk, raising the possibility of a clinically important interaction in the trials. DESIGN: The hormone trials were randomized, double-blind, and placebo-controlled. Use of nonsteroidal anti-inflammatory drugs was assessed at baseline and at years 1, 3, and 6. SETTING: The Women's Health Initiative hormone trials were conducted at 40 clinical sites in the United States. PARTICIPANTS: The trials enrolled 27,347 postmenopausal women, aged 50-79 y. INTERVENTIONS: We randomized 16,608 women with intact uterus to conjugated estrogens 0.625 mg with medroxyprogesterone acetate 2.5 mg daily or to placebo, and 10,739 women with prior hysterectomy to conjugated estrogens 0.625 mg daily or placebo. OUTCOME MEASURES: Myocardial infarction, coronary death, and coronary revascularization were ascertained during 5.6 y of follow-up in the estrogen plus progestin trial and 6.8 y of follow-up in the estrogen alone trial. RESULTS: Hazard ratios with 95% confidence intervals were calculated from Cox proportional hazard models stratified by COX inhibitor use. The hazard ratio for myocardial infarction/coronary death with estrogen plus progestin was 1.13 (95% confidence interval 0.68-1.89) among non-users of COX inhibitors, and 1.35 (95% confidence interval 0.86-2.10) among continuous users. The hazard ratio with estrogen alone was 0.92 (95% confidence interval 0.57-1.48) among non-users of COX inhibitors, and 1.08 (95% confidence interval 0.69-1.70) among continuous users. In a second analytic approach, hazard ratios were calculated from Cox models that included hormone trial assignment as well as a time-dependent covariate for medication use, and an interaction term. No significant interaction was identified. CONCLUSIONS: Use of COX inhibitors did not significantly affect the Women's Health Initiative hormone trial results

Response to comment on "Human-specific gain of function in a developmental enhancer"

Duret and Galtier argue that human-specific sequence divergence and gain of function in the HACNS1 enhancer result from deleterious biased gene conversion (BGC) with no contribution from positive selection. We reinforce our previous conclusion by analyzing hypothesized BGC events genomewide and assessing the effect of recombination rates on human-accelerated conserved noncoding sequence ascertainment. We also provide evidence that AT → GC substitution bias can coexist with positive selection

Untethered soft robotic matter with passive control of shape morphing and propulsion

There is growing interest in creating untethered soft robotic matter that can repeatedly shape-morph and self-propel in response to external stimuli. Toward this goal, we printed soft robotic matter composed of liquid crystal elastomer (LCE) bilayers with orthogonal director alignment and different nematic-to-isotropic transition temperatures (T_(NI)) to form active hinges that interconnect polymeric tiles. When heated above their respective actuation temperatures, the printed LCE hinges exhibit a large, reversible bending response. Their actuation response is programmed by varying their chemistry and printed architecture. Through an integrated design and additive manufacturing approach, we created passively controlled, untethered soft robotic matter that adopts task-specific configurations on demand, including a self-twisting origami polyhedron that exhibits three stable configurations and a “rollbot” that assembles into a pentagonal prism and self-rolls in programmed responses to thermal stimuli

Untethered soft robotic matter with passive control of shape morphing and propulsion

There is growing interest in creating untethered soft robotic matter that can repeatedly shape-morph and self-propel in response to external stimuli. Toward this goal, we printed soft robotic matter composed of liquid crystal elastomer (LCE) bilayers with orthogonal director alignment and different nematic-to-isotropic transition temperatures (T_(NI)) to form active hinges that interconnect polymeric tiles. When heated above their respective actuation temperatures, the printed LCE hinges exhibit a large, reversible bending response. Their actuation response is programmed by varying their chemistry and printed architecture. Through an integrated design and additive manufacturing approach, we created passively controlled, untethered soft robotic matter that adopts task-specific configurations on demand, including a self-twisting origami polyhedron that exhibits three stable configurations and a “rollbot” that assembles into a pentagonal prism and self-rolls in programmed responses to thermal stimuli