title: Bacterial expression and purification of the amyloidogenic peptide PAPf39 for multidimensional NMR spectroscopy.
abstract: PAPf39 is a 39 residue peptide fragment from human prostatic acidic phosphatase that forms amyloid fibrils in semen. These fibrils have been implicated in facilitating HIV transmission. To enable structural studies of PAPf39 by NMR spectroscopy, efficient methods allowing the production of milligram quantities of isotopically labeled peptide are essential. Here, we report the high-yield expression and purification of uniformly (13)C- and (15)N-labeled PAPf39 peptide, through expression as a fusion to ubiquitin at the N-terminus and an intein at the C-terminus. This allows the study of the PAPf39 monomer conformational ensemble by NMR spectroscopy. To this end, we performed the NMR chemical shift assignment of the PAPf39 peptide in the monomeric state at low pH.
authors: Center for Biotechnology and Interdisciplinary Studies and Department of Biology, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
authors: Aranganathan
authors: A
authors: Shanmuganathan
authors: Anthony C
authors: AC
authors: Bishop
authors: Kinsley C
authors: KC
authors: French
authors: Scott A
authors: SA
authors: McCallum
authors: George I
authors: GI
authors: Makhatadze
mesh_terms: D000595:Amino Acid Sequence; D000682:Amyloid / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D047668:Inteins; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D011487:Protein Conformation; D017027:Protein Tyrosine Phosphatases / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D025801:Ubiquitin / Q000235:genetics / Q000302:isolation & purification
chemical_list: D000682:Amyloid; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D025801:Ubiquitin; C526435:prostatic acid phosphatase (248-286), human; D017027:Protein Tyrosine Phosphatases