Avian H5: are humans an easy target?

Avian H5: are humans an easy target? Sinick Group: Signe Christophersen and Nicholas GauthierProtein Structure and Computational Biology, Spring 2006

Presentation Outline • Introduction to H5 • Function • Structure • H5 Model • Quality Validation • H1/H5/H9 Comparisons • Conclusions • The Uncharacterized Protein • Questions?

Introduction – H5 Function • Target cell attachment • Target receptors consist of sialic acid residues of glycosylated receptor proteins • prefers binding α2-6 in humans • prefers α2-3 linkage in avian • Release of viral contents into the cell • Lower pH in endosome causes change in HA that induces fusion of cell and virus membrane Image taken from (then modified): http://www.ch.ic.ac.uk/local/projects/sanderson/immunology.htm

Introduction – H5 Structure 1 • Homotrimer of Heterodimers • Synthesized in the cell as HA0 • Trimerizes and is transferred to cell surface

Introduction – H5 Structure 2 • Proteolytically cleaved to yield HA1 and HA2 which are held together by disulfide bonds • HA1 is shown in blue • It contains the receptor binding site which is bound to an α2-3 (avian) linkage • HA2 is shown in green • ~20 residues at the N-terminal are involved in cell/virus fusion

Models of human H5 • Has avian H5 adapted to become human specific? • Chose newest H5 isolated from human, DQ435202 • Create models using CPHmodels and HHPred • Check quality of template • Validate models • Differences between H5 found in human and avian

Quality of Template 1JSM was chosen as template for both models Resolution: 1.9Å R: 0.234 Rfree: 0.264 • PROCHECK ramachandran plot • Most favoured regions: 84.9 • Additionally allowed regions: 14.2 • Generously allowed regions: 0.5 • Disallowed regions: 0,5

Model Validation - ProQ LG > 2.5 very good model LG > 4 extremely good model MaxSub > 0.1 fairly good model MaxSub > 0.8 extremely good model Template – 1JSM CPH model Template – 1JSM HHPred model

Model Validation – VERIFY3d HA1 HA2

Receptor Binding Site HHPred model CPH model Template, 1JSM

Receptor Binding Site Differences between H5 avian and H5 human isolate 186 193 221

Receptor Binding Site 186 Av – N Hu - S 193 Av – K Hu - R CPH model Template, 1JSM

Human Adaptation How Can it Happen • Avian H1 can become human specific: E190D and G225D • Avian H3 can become human specific: Q226L and G228S • Mutations introduced into H5 (A/Vietnam/1203/2004)

Receptor Analogues

Avian H5 Receptor binding E190 Q226 G228 G225

H1 vs. H5 Receptor Binding Differences between H5 (1JSM) and H1 (1RVX)

H1 vs. H5 Receptor Binding H1 – N193 H5 – K193 H1 – D225 H5 – G225 H1 – P186 H5 – N186 H1, α2-6 receptor, H5, α2-3 receptor

H9 from swine α2-6 receptor α2-3 receptor

H9 vs. H5 Receptor Binding Differences between H5 (1JSM) and H9 (1JSD)

H9 vs. H5 Receptor Binding H9 – V190 H5 – E190 H9 – L226 H5 – Q226 H1, α2-6 H5, α2-3 H9 – P186 H5 – N186

Conclusion • H5 has not been adapted for humans • Mutations rendering H5 α2-6 specific have not been identified • We propose that human adaptation of H5 might happen via an “H9 pathway”, including the mutations E190V and Q226L. Also, the mutations N186P, L193N and G225D and might play a role. • We suggest performing mutational analysis (including these mutations), followed by analyses of receptor binding properties to gain further information of a possible human adaptation of bird flu.

Nonstructural Protein 1 • Structural model created from first 73 residues • Structural model created using CPHmodels (shown at right) • This domain is suggested to be a nucleic acid binding motif. • Last 152 residues had no structural homolog…

Nonstructural Protein 1 (cont) • Using the remaining 152 residues we… • Used HHpred to predict folds (see at right) • ProtFun: predicted to be an enzyme (65%), involvement in amino acid biosynthesis (28%) • SignalP: no signal peptide, and is non-secretory • TMHMM: found no transmembrane regions

Questions? Sinick Group: Signe Christophersen and Nicholas GauthierProtein Structure and Computational Biology, Spring 2006

Avian H5: are humans an easy target?