title: Characterizing the Structure and Oligomerization of Major Royal Jelly Protein 1 (MRJP1) by Mass Spectrometry and Complementary Biophysical Tools.
abstract: Royal jelly (RJ) triggers the development of female honeybee larvae into queens. This effect has been attributed to the presence of major royal jelly protein 1 (MRJP1) in RJ. MRJP1 isolated from royal jelly is tightly associated with apisimin, a 54-residue α-helical peptide that promotes the noncovalent assembly of MRJP1 into multimers. No high-resolution structural data are available for these complexes, and their binding stoichiometry remains uncertain. We examined MRJP1/apisimin using a range of biophysical techniques. We also investigated the behavior of deglycosylated samples, as well as samples with reduced apisimin content. Our mass spectrometry (MS) data demonstrate that the native complexes predominantly exist in a (MRJP14apisimin4) stoichiometry. Hydrogen/deuterium exchange MS reveals that MRJP1 within these complexes is extensively disordered in the range of residues 20-265. Marginally stable secondary structure (likely antiparallel β-sheet) exists around residues 266-432. These weakly structured regions interchange with conformers that are extensively unfolded, giving rise to bimodal (EX1) isotope distributions. We propose that the native complexes have a "dimer of dimers" quaternary structure in which MRJP1 chains are bridged by apisimin. Specifically, our data suggest that apisimin acts as a linker that forms hydrophobic contacts involving the MRJP1 segment316VLFFGLV322. Deglycosylation produces large soluble aggregates, highlighting the role of glycans as aggregation inhibitors. Samples with reduced apisimin content form dimeric complexes with a (MRJP12apisimin1) stoichiometry. The information uncovered in this work will help pave the way toward a better understanding of the unique physiological role played by MRJP1 during queen differentiation.
authors.affiliation: Department of Chemistry, Western University , London, Ontario, Canada N6A 5B7.|Laboratory of Biochemistry and Protein Chemistry, Department of Cell Biology, University of Brasilia , Brasilia, Brazil.
authors.forename: Samuel C
authors.initials: SC
authors.lastname: Mandacaru
authors.affiliation: Laboratory of Biochemistry and Protein Chemistry, Department of Cell Biology, University of Brasilia , Brasilia, Brazil.|Proteomics Center of Excellence, Departments of Chemistry and Molecular Biosciences, Northwestern University , Evanston, Illinois 60611, United States.
authors.forename: Luis H F
authors.initials: LH
authors.lastname: do Vale
authors.affiliation: Department of Chemistry, Western University , London, Ontario, Canada N6A 5B7.
authors.forename: Siavash
authors.initials: S
authors.lastname: Vahidi
authors.affiliation: Department of Chemistry, Western University , London, Ontario, Canada N6A 5B7.
authors.forename: Yiming
authors.initials: Y
authors.lastname: Xiao
authors.affiliation: Proteomics Center of Excellence, Departments of Chemistry and Molecular Biosciences, Northwestern University , Evanston, Illinois 60611, United States.
authors.forename: Owen S
authors.initials: OS
authors.lastname: Skinner
authors.affiliation: Laboratory of Biochemistry and Protein Chemistry, Department of Cell Biology, University of Brasilia , Brasilia, Brazil.
authors.forename: Carlos A O
authors.initials: CA
authors.lastname: Ricart
authors.affiliation: Proteomics Center of Excellence, Departments of Chemistry and Molecular Biosciences, Northwestern University , Evanston, Illinois 60611, United States.
authors.forename: Neil L
authors.identifier: 0000-0002-8815-3372
authors.initials: NL
authors.lastname: Kelleher
authors.affiliation: Laboratory of Biochemistry and Protein Chemistry, Department of Cell Biology, University of Brasilia , Brasilia, Brazil.
authors.forename: Marcelo Valle
authors.initials: MV
authors.lastname: de Sousa
authors.affiliation: Department of Chemistry, Western University , London, Ontario, Canada N6A 5B7.
authors.forename: Lars
authors.identifier: 0000-0002-5283-3165
authors.initials: L
authors.lastname: Konermann
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001516:Bees / Q000254:growth & development / Q000378:metabolism; D041961:Deuterium Exchange Measurement; D005227:Fatty Acids / Q000737:chemistry* / Q000502:physiology; D015870:Gene Expression; D006023:Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019476:Insect Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D064267:Intrinsically Disordered Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007814:Larva / Q000254:growth & development / Q000378:metabolism; D013058:Mass Spectrometry; D018832:Molecular Chaperones / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011134:Polysaccharides / Q000737:chemistry* / Q000378:metabolism; D055503:Protein Multimerization; D000099219:Royal Jelly
chemical_list: D005227:Fatty Acids; D006023:Glycoproteins; D019476:Insect Proteins; D064267:Intrinsically Disordered Proteins; D018832:Molecular Chaperones; D010455:Peptides; D011134:Polysaccharides; D000099219:Royal Jelly; C559157:MRJP1 protein, Apis mellifera