Mussel foot protein-3 inMytilus californianus musseladhesive plaquesMiguel JassoMentor: Nadine Martinez RodriguezDepartment: Molecular, Cellular, andDevelopmental Biology (MCDB)Advisors: Herbert Waite and Jacob Israelachvili
Mussels and their Plaques• Attach to surfacesunderwater.• Threads extend from themussel with adhesiveplaques• Adhesive plaques aremade up of 6+ differentproteins.• Study properties todevelop under wateradhesivesMytilus californianus
Distribution of mytilus californianus footproteins (mcfp)Proposed diagram of mussel plaque• Mcfp-3• Interface• Each mcfp has adifferent role.Source: Hwang, D.S. et. al., 2007Goal: Investigate the distribution of the mcfp-3 in the plaque.
Overall Procedure for Locatingmcfp-3Collect plaques frommussels and embedthem into resin.Use a microtome tosection plaques intodifferent layers.Representative figure of a MALDI-TOF mass spectrumPic Source: www.bioacademy.gr/bioinformatics/Analyze layers withMALDI-TOF todetermine thelocation of mcfp-3
Mussel Plaque Harvesting• Mussels attach to plastic plates.• Plaques are separated from threads.• Plaques placed in vials of seawater.
Embedding the PlaquesplaquesSoaked in PBS/EtOHSoaked in milli-Q H2O/ acetoneSoaked in resinTransfer tomoldsBaked at 70°C
Microtome• Sections samples withprecision• Plaques are sectioned• Placed on MALDI goldplateDirection of plaque slicing
MALDI-TOFPic Source: protein.iastate.edu
Unexpected Results• Did not fly at all in the MALDI-TOF• Resin did not allow the proteins to fly• Dehydration steps fixed proteins• Plaques aged• Modified technique• Used fresh whole plaques• Dissection into three parts
Modified Sectioning of Plaques• Plaques sectionedwith clean single edgerazor• Applied to gold plate• Subjected to MALDI-TOF MSSource: Hwang, D.S. et. al., 2007
Mcfp-3 and -6 are found at thejunction between the thread andplaque•Mcfp-3 ~ 5.5 kDa•Mcfp-6 ~ 11 kDamcfp-3: 5586.08 mcfp-6: 11706.4Source: Hwang, D.S. et. al., 2007
Mcfp-3 and -6 are found in the middleof the plaque•Mcfp-3 ~ 5.5 kDa•Mcfp-6 ~ 11 kDamcfp-3:5295.155546.75mcfp-6:11571.6Source: Hwang, D.S. et. al., 2007
Mcfp-3 and -6 are located at theinterface of the plaque• Mcfp-3 ~ 5.5 kDa• Mcfp-6 ~ 11kDamcfp-3:5509.025645.18mcfp-6:11495.2Source: Hwang, D.S. et. al., 2007
Mcfp-3 and -6 are located near theperimeter of the interface•Mcfp-3 ~ 5.5 kDa•Mcfp-6 ~ 11 kDamcfp-3: 5275.14mcfp-6: 11509.3Source: Hwang, D.S. et. al., 2007
Conclusion• The location of mcfp-3 at the interface is inagreement with previous published results.• Gives insight to how the proteins interactthroughout the plaque.• These results suggest that mcfp-3 is locatedthrough out the plaque.
Reflections• Hands on experience with working a lab.• Appreciation for marine biotechnology• Consider in future plans• The AR program really influenced me topursue a science career.
Acknowledgements• I would like to say thanks to:• My mentor Nadine Martinez Rodriguez• Dr. Waite• Lubi Lenaburg• Chris Hammetter• Alison Butler• Julia Callender• Other mentors, student and interns in my lab• California Nano Systems Institute (CNSI),Center for Science and EngineeringPartnerships (CSEP), LEAPS,