Biology subjects

Drew Endy

Publications and source records attributed to Drew Endy.

4 recordsLinked to original sources

bioRxiv2016

Measurements of translation initiation from all 64 codons in E. coli

Our understanding of translation is one cornerstone of molecular biology that underpins our capacity to engineer living matter. The canonical start codon (AUG) and a few near-cognates (GUG, UUG) are typically considered as the \"start codons\" for translation initiation in Escherichia coli (E. coli). Translation is typically not thought to initiate from the 61 remaining codons. Here, we systematically quantified translation initiation in E. coli from all 64 triplet codons. We detected protein synthesis above background initiating from at least 46 codons. Translation initiated from these non-canonical start codons at levels ranging from 0.01% to 2% relative to AUG. Translation initiation from non-canonical start codons may contribute to the synthesis of peptides in both natural and synthetic biological systems

Synthetic Biology↗

bioRxiv2016

When Wavelengths Collide: Bias in Cell Abundance Measurements due to Expressed Fluorescent Proteins

The abundance of bacteria in liquid culture is commonly inferred by measuring optical density at 600 nm. Red fluorescent proteins (RFP) can strongly absorb light at 600 nm. Increasing RFP expression can falsely inflate apparent cell density and lead to underestimations of mean per-cell fluorescence by up to 10%. Measuring optical density at 700 nm would allow estimation of cell abundance unaffected by the presence of nearly all fluorescent proteins.

Synthetic Biology↗

bioRxiv2015

Connecting Boolean Integrase Logic gates to a novel alkane control signal via engineered level matching.

Boolean Integrase Logic gates use phage integrases that respond to transcriptional signals to implement logical functions within living cells via DNA recombination. Control of biological systems using these logic gates has many applications, including biomanufacturing, healthcare, or environmental remediation. Gates are highly sensitive to background transcriptional noise producing unwanted integrase expression and uncontrolled, permanent gate switching. Consequently, connecting gates to novel control signals requires time-consuming directed evolution of regions regulating integrase expression. Here we present an approach in which the activity of an alkane biosensor is tuned to match levels of existing integrase control signals. By adjusting AlkS expression, we tuned the transcriptional output of Palkb to match the output of existing integrase controllers. We successfully connected Palkb to two integrases with different transcriptional control requirements and demonstrated the original logic function was conserved. Our method complements directed evolution approaches to connect Boolean integrate logic gates to novel transcriptional sources and will ultimately facilitate the systematic in silico design of gates responding to various control signals.

Synthetic Biology↗

bioRxiv2015

Complete absence of thebaine biosynthesis under home-brew fermentation conditions

Yeast-based biosynthesis of medicinal compounds traditionally derived from plant materials is improving. Both concerns and hopes exist for the possibility that individual small volume batch fermentations could provide distributed and independent access to a diversity of compounds some of which are now abused, illegal, or unavailable to many who need for genuine medical purposes. However, there are differences between industrial bioreactors and home-brew fermentation. We used engineered yeast that make thebaine, a morphinan opiate, to quantify if differences in fermentation conditions impact biosynthesis yields. We used yeast that make an English ale as a positive fermentation control. We observed no production of thebaine and miniscule amounts of reticuline, an upstream biosynthetic intermediate, in home-brew fermentations; the positive control was palatable. We suggest that additional technical challenges, some of which are unknown and likely unrelated to optimized production in large-volume bioreactors, would need to be addressed for engineered yeast to ever realize home-brew biosynthesis of medicinal opiates at meaningful yields.

Synthetic Biology↗