MSc Thesis Presentation: Anabaena Sensory Rhodopsin reconstituted in nanodiscs as a promising platform for structural studies

MSc Candidate

Suelen Camargo

Abstract

Membrane protein studies need an excellent membrane-mimicking reconstitution system. Nanodiscs offer many advantages in this regard but require optimal conditions for achieving homogeneous and stable samples. Model protein Anabaena Sensory Rhodopsin (ASR) characterization in nanodiscs provides insights into protocol steps of membrane proteins reconstitution. Size-exclusion chromatography and electron microscopy aided in sample evaluation, identifying aggregation and the disk diameter, making possible estimation of ASR oligomeric state. Solution-state NMR showed ASR stability in nanodiscs for at least five days. Comparing liposomes and nanodiscs via ssNMR showed that ASR structure is mostly unaffected, minimally impacting loops. Helical arrangement alters somewhat, due to lipid composition and MSP scaffold influence, possibly causing direct MSP-ASR interaction. ssNMR detected co-purified PE lipids but not ECAPG, suggesting that ASR in nanodiscs might lack these sugar moieties due to structural changes. This research contributes to an improved comprehension of ASR's structure and oligomerization behavior across varied reconstitution platforms.

Examination Committee

• Dr. Michele Oliver, Chair
• Dr. Leonid Brown, Advisor
• Dr. Vladimir Ladizhansky Co-Advisor
• Dr. Robert Wickham, Graduate Faculty