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NIH grant 1R01GM165765-01: Optical Probes for Polymerase Engineering
Institute: {'code': 'GM', 'abbreviation': 'NIGMS', 'name': 'National Institute of General Medical Sciences', 'admin_org_id': '12500024', 'admin_funding_url': ''}. PI: John Charles Chaput. Award: $1188418.
Start: 2026-09-05T00:00:00.
PROJECT SUMMARY/ABSTRACT
Polymerases are among the most powerful tools in the molecular biology toolbox, allowing researchers to
precisely synthesize DNA and RNA molecules for diverse applications in synthetic biology and healthcare.
But what if these enzymes were no longer restricted to the genetic polymers of life? What new technologies
could we develop, or what new discoveries could we make? One clear example is the use of specialty
polymerases to support the evolution of affinity reagents (aptamers) and catalysts that are better equipped
to function under harsh biological conditions. Motivated by this goal, examples of engineered polymerases
have emerged that can synthesize diverse classes of genetic molecules (XNAs). However, despite many
impressive advances, the widespread adoption of XNAs in biomedical applications remains limited by the
low activity of existing polymerases. The central challenge lies in understanding how to reshape the active
site pocket to accommodate highly divergent substrates while preserving the catalytic activity of the native
enzyme fold. Fortunately, recent advances in high-throughput microfluidic screening platforms, like droplet-
based optical polymerase sorting (DrOPS), provide hope that a solution is finally within reach.
This proposal seeks to overcome the remaining technical barriers needed to advance DrOPS as a versatile
platform for engineering polymerases with custom activities. Unlike conventional approaches that require
days to screen 103-104 library members, DrOPS can query 108 members per day using 106-fold less
sample volume. Its favorable cost efficiency and minimal selection bias make it well-suited for polymerase
engineering. Recognizing these benefits, the proposed study will provide a more accessible and enabling
technology for polymerase engineering by advancing the design of molecular probes that will function in
uniform microdroplets as optical sensors of polymerase activity. The probes, once designed and tested,
will be validated in directed evolution experiments aimed at producing new types of XNA polymerases that
can be used as research tools for drug discovery. Together, these goals will expand and refine the DrOPS
technology platform from a working prototype to a highly advanced system that can address challenging
problems in enzyme engineering.document fetched 2026-09-13 02:45