Poster Title

Visualization of an Ineffective T-State Stabilizing Mutation in Circularly Permuted Hemoglobin

Co-Author(s)

Johann Sigurjonsson, P. Clint Spiegel, Spencer Anthony-Cahill

Research Mentor(s)

Spencer Anthony-Cahill

Affiliated Department

Chemistry

Sort Order

66

Start Date

18-5-2017 9:00 AM

End Date

18-5-2017 12:00 PM

Document Type

Event

Abstract

The primary goal of our project is to develop a functional hemoglobin based oxygen carrier (HBOC) for clinical care. In order to accomplish this, we are working on designing a single chain hemoglobin (scHb) that can serve as the building block of a monodisperse polymeric HBOC. The scHb is formed from novel covalent fusions between the two α chains and two circularly permuted β chains. The initial constructs of non-covalently linked α and cp-β showed decreased stability and low T-state affinity (Asmundson, et. al., 2009). To counter these effects, a point mutation was included to increase T-state affinity (Tsai et. al in 1999). However, there is evidence that this mutation, αV96W, may in fact further decrease T-state affinity (Apperson, J., 2013). We have initiated X-ray crystallography trials of αV96W mutants to understand the structural basis for this unexpected effect on the T-state stability, to inform future protein engineering of these HBOC candidates.

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 documentation for commercial purposes, or for financial gain, shall not be allowed without the author's written permission.

Language

English

Format

application/pdf

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May 18th, 9:00 AM May 18th, 12:00 PM

Visualization of an Ineffective T-State Stabilizing Mutation in Circularly Permuted Hemoglobin

Chemistry

The primary goal of our project is to develop a functional hemoglobin based oxygen carrier (HBOC) for clinical care. In order to accomplish this, we are working on designing a single chain hemoglobin (scHb) that can serve as the building block of a monodisperse polymeric HBOC. The scHb is formed from novel covalent fusions between the two α chains and two circularly permuted β chains. The initial constructs of non-covalently linked α and cp-β showed decreased stability and low T-state affinity (Asmundson, et. al., 2009). To counter these effects, a point mutation was included to increase T-state affinity (Tsai et. al in 1999). However, there is evidence that this mutation, αV96W, may in fact further decrease T-state affinity (Apperson, J., 2013). We have initiated X-ray crystallography trials of αV96W mutants to understand the structural basis for this unexpected effect on the T-state stability, to inform future protein engineering of these HBOC candidates.