Improved subtiligase variants allow broad and versatile site-specific chemical modification or conjugation of proteins on their N-termini.
Site-specific chemical modification of proteins is an enabling technology for drug design, chemical biology, cell biology, and materials science. The existing subtiligase technology selectively labels proteins on their N-termini with synthetic peptides. However, many of the N-terminal dipeptide sequences were inaccessible to subtiligase labeling due to incompatible sequences. Investigators at UCSF have made a significant improvement over current technology by engineering new subtiligase variants that allow bioconjugation to many more different N-terminal sequences and accept various peptide ester substrates. Notably, these include sequences that have an acidic residue at the N terminus, which is often the case for mammalian antibodies and therefore allows application in the production of antibody-drug conjugates.
The Wells Lab at UCSF has engineered new variants of subtiligase with altered N-terminal protein substrate specificity to enable broadened site-specific protein bioconjugation. These variants also have an increased or improved aminolysis to hydrolysis ratio (A/H ratio) compared to a wild-type subtiligase or a wild-type stabiligase. The use of these novel enzymes allows selective protein labeling on many more N-terminal sequences than were previously accessible to subtiligase labeling. The inventors have also developed an algorithm that facilitates the selection of appropriate subtiligase mutants for the N-terminal sequences of interest. Such site-specific bioconjugation can be used to link polypeptides to fluorophores, affinity handles, DNA barcodes, drug payloads, and reactive groups.
To develop & commercialize the subtiligase variants and their substrates as a kit or tool for site-specific protein conjugation to use in antibody-drug conjugation, drug design, and as a research tool for imaging, fluorescence assays, proteomic profiling and affinity pulldowns.
Proof of concept
Under CDA / NDA
|Patent Cooperation Treaty||Published Application||WO2018045200||03/08/2018||2017-035|
Additional Patent Pending
Site-Specific Protein Labeling, Subtiligase, N-termini, Antibody-Drug Conjugation (ADC), Protein Profiling, Research Tools, Proteomics