Poster Annual Conference of the Genetics Society of Australasia with the NZ Society for Biochemistry & Molecular Biology

Understanding the Batten disease associated protein CLN5 (666)

Jade JR Yip 1 2 3 , Hollie E Wicky 1 2 3 4 , Hannah L Best 1 2 3 , Peter D Mace 1 , Stephanie M Hughes 1 2 3 4
  1. Department of Biochemistry, University of Otago, Dunedin, New Zealand
  2. Brain Health Research Centre, University of Otago, Dunedin, New Zealand
  3. Genetics Otago, University of Otago, Dunedin, New Zealand
  4. Brain Research New Zealand, University of Otago, Dunedin, New Zealand

The neuronal ceroid lipofuscinoses (NCL, Batten disease) are a group of autosomal recessive neurodegenerative lysosomal storage diseases that typically result in loss of vision, epilepsy, loss of motor function and cognitive decline. Mutations throughout the CLN5 gene predominantly cause a late-infantile variant of NCL. The CLN5 protein is a soluble lysosomal protein of unknown function, with no strong sequence identity to any previously characterised protein. Here we aim to better understand the normal role of the CLN5 protein by investigating its structure and potential function. Using secondary structure prediction, sequence profile matching, and homology modelling, we have found that CLN5 shares weak homology with proteases that utilise a cysteine histidine catalytic dyad. Based on this, the predicted catalytic cysteine was mutated via site directed mutagenesis to investigate whether this mutant can rescue disease-associated phenotypes present in CLN5-/- affected cells. Prior to testing for phenotypic rescue, we will compare the trafficking and secretion of mutant and wildtype CLN5. 

Complementary to functional studies, an optimised expression construct for CLN5 has been designed for purification and crystallisation studies. HEK293FT cells have been modified using lentiviral transduction to overexpress CLN5. CLN5 protein has been collected from the media of these cells and purified using nickel affinity chromatography. Future work is aimed at determining the structure of CLN5.

Combined, this work will lead to a better understanding of the normal function of CLN5, and as to why mutations in the gene lead to NCL.