Generation of Polymeric Recombinant Hemoglobin Using DBCO-modified Hemoglobin
Research Mentor(s)
Spencer Anthony-Cahil
Description
Oligomerized hemoglobin (Hb) molecules have been shown to decrease previously observed deleterious effects from treatment with cell-free hemoglobin1. To create these poly-Hb constructs, a method has developed which employs the site specific ligation reaction of the sortase A enzyme from S. aureus. Hb mutants generated in our lab (αcpβ) have been further modified by adding the sortase recognition sequence, LPXTG, to the C-terminus of the α-subunit (s-αcpβ). Sortase-mediated ligation (SML) will be employed to ligate Hb subunits to a peptide linker that contains a strained cyclooctyne (DBCO), which in turn we plan to react with scaffolding molecules that display azides. We propose to control the final oligomerization state of the poly-Hb by the structure of the scaffold, which will have a defined number of reactive azides that can form covalent bonds with the DBCO-modified Hbs.
Document Type
Event
Start Date
15-5-2019 9:00 AM
End Date
15-5-2019 5:00 PM
Location
Carver Gym (Bellingham, Wash.)
Department
Chemistry
Genre/Form
student projects, posters
Subjects – Topical (LCSH)
Hemoglobin--Synthesis; Oligomers--Synthesis; Ligases; Recombinant DNA
Type
Image
Rights
Copying of this document in whole or in part is allowable only for scholarly purposes. It is understood, however, that any copying or publication of this document for commercial purposes, or for financial gain, shall not be allowed without the author’s written permission.
Language
English
Format
application/pdf
Generation of Polymeric Recombinant Hemoglobin Using DBCO-modified Hemoglobin
Carver Gym (Bellingham, Wash.)
Oligomerized hemoglobin (Hb) molecules have been shown to decrease previously observed deleterious effects from treatment with cell-free hemoglobin1. To create these poly-Hb constructs, a method has developed which employs the site specific ligation reaction of the sortase A enzyme from S. aureus. Hb mutants generated in our lab (αcpβ) have been further modified by adding the sortase recognition sequence, LPXTG, to the C-terminus of the α-subunit (s-αcpβ). Sortase-mediated ligation (SML) will be employed to ligate Hb subunits to a peptide linker that contains a strained cyclooctyne (DBCO), which in turn we plan to react with scaffolding molecules that display azides. We propose to control the final oligomerization state of the poly-Hb by the structure of the scaffold, which will have a defined number of reactive azides that can form covalent bonds with the DBCO-modified Hbs.