Fab MOR03268 triggers absorption shift of a diagnostic dye via packaging in a solvent-shielded Fab dimer interface.
Molecular interactions between near-IR fluorescent probes and specific antibodies may be exploited to generate novel smart probes for diagnostic imaging. Using a new phage display technology, we developed such antibody Fab fragments with subnanomolar binding affinity for tetrasulfocyanine, a near-IR in vivo imaging agent. Unexpectedly, some Fabs induced redshifts of the dye absorption peak of up to 44 nm. This is the largest shift reported for a biological system so far. Crystal structure determination and absorption spectroscopy in the crystal in combination with microcalorimetry and small-angle X-ray scattering in solution revealed that the redshift is triggered by formation of a Fab dimer, with tetrasulfocyanine being buried in a fully closed protein cavity within the dimer interface. The derived principle of shifting the absorption peak of a symmetric dye via packaging within a Fab dimer interface may be transferred to other diagnostic fluorophores, opening the way towards smart imaging probes that change their wavelength upon interaction with an antibody.
Hillig
Roman C
RC
Bayer Schering Pharma AG, Global Drug Discovery, 13342 Berlin, Germany. roman.hillig@bayerhealthcare.com
Urlinger
Stefanie
S
Fanghänel
Jörg
J
Brocks
Bodo
B
Haenel
Cornelia
C
Stark
Yvonne
Y
Sülzle
Detlev
D
Svergun
Dmitri I
DI
Baesler
Siegfried
S
Malawski
Guido
G
Moosmayer
Dieter
D
Menrad
Andreas
A
Schirner
Michael
M
Licha
Kai
K
D000042:Absorption; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology*; D000915:Antibody Affinity; D002151:Calorimetry; D002850:Chromatography, Gel; D004396:Coloring Agents / Q000737:chemistry*; D022801:Complementarity Determining Regions / Q000737:chemistry; D018360:Crystallography, X-Ray; D019281:Dimerization; D005453:Fluorescence; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry* / Q000276:immunology*; D007211:Indoles / Q000737:chemistry* / Q000276:immunology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019151:Peptide Library; D053838:Scattering, Small Angle; D012997:Solvents / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet
D000911:Antibodies, Monoclonal; D004396:Coloring Agents; D022801:Complementarity Determining Regions; C527478:Fab MOR03268; D007140:Immunoglobulin Fab Fragments; D007211:Indoles; D019151:Peptide Library; D012997:Solvents