Novel affinity tag system using structurally defined antibody-tag interaction: application to single-step protein purification.
Biologically important human proteins often require mammalian cell expression for structural studies, presenting technical and economical problems in the production/purification processes. We introduce a novel affinity peptide tagging system that uses a low affinity anti-peptide monoclonal antibody. Concatenation of the short recognition sequence enabled the successful engineering of an 18-residue affinity tag with ideal solution binding kinetics, providing a low-cost purification means when combined with nondenaturing elution by water-miscible organic solvents. Three-dimensional information provides a firm structural basis for the antibody-peptide interaction, opening opportunities for further improvements/modifications.
Nogi
Terukazu
T
Laboratory of Protein Synthesis and Expression, Institute for Protein Research, Osaka University, Suita, Osaka 565-0871, Japan.
Sangawa
Takeshi
T
Tabata
Sanae
S
Nagae
Masamichi
M
Tamura-Kawakami
Keiko
K
Beppu
Ayako
A
Hattori
Mitsuharu
M
Yasui
Norihisa
N
Takagi
Junichi
J
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D000915:Antibody Affinity; D015816:Cell Adhesion Molecules, Neuronal / Q000235:genetics / Q000302:isolation & purification; D045744:Cell Line, Tumor; D002846:Chromatography, Affinity / Q000379:methods; D018360:Crystallography, X-Ray / Q000379:methods; D000939:Epitopes / Q000378:metabolism*; D016326:Extracellular Matrix Proteins / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D007700:Kinetics; D051379:Mice; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000302:isolation & purification; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D011485:Protein Binding; D011506:Proteins / Q000235:genetics / Q000302:isolation & purification*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D000091042:Reelin Protein; D012697:Serine Endopeptidases / Q000235:genetics / Q000302:isolation & purification; D013329:Structure-Activity Relationship
D000911:Antibodies, Monoclonal; D015816:Cell Adhesion Molecules, Neuronal; D000939:Epitopes; D016326:Extracellular Matrix Proteins; D007140:Immunoglobulin Fab Fragments; D009419:Nerve Tissue Proteins; D010455:Peptides; D011506:Proteins; D011993:Recombinant Fusion Proteins; D000091042:Reelin Protein; C429104:SPON1 protein, human; D012697:Serine Endopeptidases
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