1. Neurotrophins & NTs Receptors Loredana Lombardi Ariadna Laguna Molecular Mechanisms of Development (EMBO), Barcelona 2006

2. Neurotrophins : Ligands, Receptors and their pleiotropic role in Development Cell DeathCell Survival, Growth and Differentiation

3. Aim Analyse the sequences of neurotrophins and their receptors using bioinformatic tools to better understand their biological properties. Procedure 1)Retrieve protein sequences from databases (mouse) 2)Align protein sequences 3)Search for known protein domains 4)Search for post-translational modifications in NTs 5)Search for phosphorylation sites and possible binding partners in Trks

4. 1)Retrieve protein sequences from databases (mouse) NameNCBISwissProtLengthpI/MW Nerve Growth Factor (NGF) AAA37686.1P01139307 aa9.56 / 27 kDa Brain Derived Neurotrophic Factor (BDNF) NP_031566P21237249 aa8.91 / 28 kDa Neurotrophin 3 (NT3) NP_032768P20181258 aa9.24 / 29 kDa Neurotrophin 4/5 (NT4/5) NP_937833209 aa9.17 / 22 kDa NameNCBISwissProtLengthpI/MW Tyrosine Receptor Kinase A (TrkA) XP_283871799 aa5.87 / 88 kDa Tyrosine Receptor Kinase B (TrkB) NP_001020245P15209821 aa6.08 / 92 kDa Tyrosine Receptor Kinase C (TrkC) NP_032772825 aa6.20 / 93 kDa p75 NP_150086427 aa4.60 / 45 kDa

5. 2-3) Align protein sequences and search for known protein domains Clustalw, Multialign, Boxshade Prosite, Pfam, Motifscan,SMART Signal Peptide Propeptide NGF domain Glycosilation (Asparagine) Disulfid Bonds

6. 2-3) Analyse p75 protein sequence and search for known protein domains Prosite, Pfam, Motifscan,SMART Signal Peptide Trans Membrane Domain DEATH Domain 4 TNFR Domains TMPRED PSIPRED

7. 2-3) Align protein sequences and search for known protein domains Clustalw, Multialign, Boxshade Prosite, Pfam, Motifscan,SMART Signal Peptide IgG like domain Leucine Rich

8. 2-3) Align protein sequences and search for known protein domains Clustalw, Multialign, Boxshade Prosite, Pfam, Motifscan,SMART Predicted P-Tyr TransMembrane Domain Tyrosine Kinase Domain TMPRED PSIPRED NetPhos 2.0 Server ATP binding site Activation Loop

9. 5) Search for phosphorylation sites and possible binding partners in Trks SCANSITE TrkA

10. Conclusions We found some discrepancies between different servers in the predicted protein domains  essential to compare different predictions !!! We found a good correspondance between different servers in the most conserved domains  NGF domain in NT and PTyrK domain in Trks It is useful to predict post-translational modifications and phosphorylation sites with bioinformatic tools to later manipulate the protein in the lab.