8 research outputs found
Building biosecurity for synthetic biology.
The fast-paced field of synthetic biology is fundamentally changing the global biosecurity framework. Current biosecurity regulations and strategies are based on previous governance paradigms for pathogen-oriented security, recombinant DNA research, and broader concerns related to genetically modified organisms (GMOs). Many scholarly discussions and biosecurity practitioners are therefore concerned that synthetic biology outpaces established biosafety and biosecurity measures to prevent deliberate and malicious or inadvertent and accidental misuse of synthetic biology's processes or products. This commentary proposes three strategies to improve biosecurity: Security must be treated as an investment in the future applicability of the technology; social scientists and policy makers should be engaged early in technology development and forecasting; and coordination among global stakeholders is necessary to ensure acceptable levels of risk
Prevalent, protective, and convergent IgG recognition of SARS-CoV-2 non-RBD spike epitopes
The molecular composition and binding epitopes of the immunoglobulin G (IgG) antibodies that circulate in blood plasma following SARS-CoV-2 infection are unknown. Proteomic deconvolution of the IgG repertoire to the spike glycoprotein in convalescent subjects revealed that the response is directed predominantly (>80%) against epitopes residing outside the receptor-binding domain (RBD). In one subject, just four IgG lineages accounted for 93.5% of the response, including an N-terminal domain (NTD)-directed antibody that was protective against lethal viral challenge. Genetic, structural, and functional characterization of a multi-donor class of âpublicâ antibodies revealed an NTD epitope that is recurrently mutated among emerging SARS-CoV-2 variants of concern. These data show that âpublicâ NTD-directed and other non-RBD plasma antibodies are prevalent and have implications for SARS-CoV-2 protection and antibody escape
Cellular reagents for diagnostics and synthetic biology.
We have found that the overproduction of enzymes in bacteria followed by their lyophilization leads to 'cellular reagents' that can be directly used to carry out numerous molecular biology reactions. We demonstrate the use of cellular reagents in a variety of molecular diagnostics, such as TaqMan qPCR with no diminution in sensitivity, and in synthetic biology cornerstones such as the Gibson assembly of DNA fragments, where new plasmids can be constructed solely based on adding cellular reagents. Cellular reagents have significantly reduced complexity and cost of production, storage and implementation, features that should facilitate accessibility and use in resource-poor conditions
Charge Shielding Prevents Aggregation of Supercharged GFP Variants at High Protein Concentration
Understanding protein stability is
central to combatting protein
aggregation diseases and developing new protein therapeutics. At the
high concentrations often present in biological systems, purified
proteins can exhibit undesirable high solution viscosities and poor
solubilities mediated by short-range electrostatic and hydrophobic
proteinâprotein interactions. The interplay between protein
amino acid sequence, protein structure, and solvent conditions to
minimize proteinâprotein interactions is key to designing well-behaved
pharmaceutical proteins. However, theoretical approaches have yet
to yield a general framework to address these problems. Here, we analyzed
the high concentration behavior of superfolder GFP (sfGFP) and two
supercharged sfGFP variants engineered to have formal charges of â18
or +15. Under low cosolute conditions, sfGFP and the â18 variant
formed a gel or phase separated at âŒ10 mg/mL. Under conditions
that screen surface charges, including formulations with high histidine
or high NaCl concentrations, all three variants attained concentrations
up to 250 mg/mL with moderate viscosities. Moreover, all three variants
exhibited very similar viscosityâconcentration profiles over
this range. This effect was not mimicked by high sugar concentrations
that exert excluded-volume effects without shielding charge. Collectively,
these data demonstrate that charge shielding neutralizes not only
long-range electrostatic interactions but also, surprisingly, short-range
electrostatic effects due to surface charge anisotropy. This work
shows that supercharged sfGFP behavior under high ionic strength is
largely determined by particle geometry, a conclusion that is supported
by colloid models and may be applicable to pharmaceutically relevant
proteins