orals 5th Asia-Pacific NMR Symposium 2013

Flagella as a Novel Alignment Media for the Measurement of Residual Dipolar Couplings in Proteins (#68)

Kandala V.R Chary 1 2 , Himanshu Singh 1 , Manish Shukla 1 , Basuthkar J Rao 1
  1. Tata Institute of Fundamental Research, Mumbai, India
  2. Tata Institute of Fundamental Research, Center for Interdisciplinary Sciences, Hyderabad, India

We demonstrate for the first time the potential use of the flagella (flexible rod-like structures; ~500 nm in diameter and 5-15 µm long) of Chlamydomonas reinhardtii, a unicellular green alga of about 10 µm in diameter, as a novel alignment media that can weakly align molecules. This media enabled the measurement of residual dipolar couplings, over a wide range of temperature and pH and even in the presence of high ionic strength or in the presence of detergents, using solution NMR spectroscopy. We have detached these flagella from C. reinhardtii by adopting low-pH sucrose method and purified them from cellular debris. This method and its reimplementation following regeneration of flagella from the same deflagellated cells provide an economical and continual supply of flagella. The flagella, being stable biological products, are highly consistent in their quality across various preparations and induce a very significant alignment order quiet comparable with other alignment media.