Sensor for Nadp (h) and Development of Alcohol Dehydrogenases

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This invention discloses a novel NADP(H) nanosensor system that uses a genetically-encoded regulatory sequence and promoter responsive to the redox state of NADP(H) in a cell. The sensor triggers expression of an autofluorescent protein, so the cell's fluorescence level provides a direct, non-invasive readout of NADP(H) availability and, by extension, enzyme activity dependent on NADP(H). This system can be engineered into cells to rapidly screen and isolate genes encoding for NADP(H)-dependent enzymes (like alcohol dehydrogenases), enabling high-throughput discovery and evolution of such enzymes without the need for laborious product measurement or cell lysis.

Use CasesContent extracted from patent full text and abstract with AI.

• High-throughput screening for NADP(H)-dependent enzymes in biotechnology and pharmaceutical research.
• Directed evolution and engineering of enzymes (such as alcohol dehydrogenases) for improved industrial biocatalysis.
• Selecting microbial strains for efficient biosynthetic production of pharmaceuticals or chemicals.
• Rapid, in vivo detection of enzyme activity profiles in synthetic biology and metabolic engineering workflows.
• Screening gene libraries from metagenomic DNA for novel redox enzymes.

BenefitsContent extracted from patent full text and abstract with AI.

• Allows simple, real-time, and non-destructive detection of NADP(H)-dependent enzyme activity inside living cells.
• Enables high-throughput screening and sorting of cells using fluorescence, dramatically accelerating enzyme discovery and optimization.
• Reduces or eliminates the need for complex, time-consuming analytical chemistry or cell lysis steps.
• Facilitates direct identification and isolation of genes encoding desirable enzymes from large gene libraries.
• Can improve the efficiency of developing bio-based manufacturing processes by enabling faster enzyme evolution and microbial strain selection.

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Patent Abstract

The invention relates to an NADP(H)-nanosensor, comprising i) a nucleic acid sequence, to which a regulator can bind, wherein the oxidation state of the regulator is dependent on the NADP(H)-availability; ii) a promoter sequence that follows the nucleic acid sequence i), to which a RNA-polymerase can bind, wherein the affinity of the RNA-polymerase for the promoter sequence is influenced by the oxidation state of the regulator; iii) a nucleic acid sequence being under the control of the promoter sequence ii), which nucleic acid sequence encodes for an auto fluorescence protein. The invention further relates to a cell, a method for isolating genes, which encode for NADP(H)-dependent enzymes, as well as the use of an NADP (H) nanosensor.