Nanosecond Motions Of Genetically-Engineered Antibodies: Structural Elements Controlling Segmental Flexibility Defined By Time-Resolved Emission Anisotropy

Theodore G Wensel; William P. Schneider; Vernon T Oi; Lubert Stryel

doi:10.1117/12.945374

24 June 1988 Nanosecond Motions Of Genetically-Engineered Antibodies: Structural Elements Controlling Segmental Flexibility Defined By Time-Resolved Emission Anisotropy

Theodore G Wensel, William P. Schneider, Vernon T Oi, Lubert Stryel

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Proceedings Volume 0909, Time-Resolved Laser Spectroscopy in Biochemistry; (1988) https://doi.org/10.1117/12.945374
Event: 1988 Los Angeles Symposium: O-E/LASE '88, 1988, Los Angeles, CA, United States

Abstract

Immunoglobulins are flexible proteins which display large-amplitude modes of motion on a nanosecond time scale. Different classes of antibodies differ markedly in their degree of segmental flexibility; a number of their essential biological functions are correlated with their nanosecond internal dynamics. These motions can be conveniently monitored by time-resolved fluorescence anisotropy measurements. An instrument built around a synch-pumped cavity-dumped dye laser and a fast time-to-digital convertor with histogramming memory has made it possible to obtain high-quality anisotropy in a few minutes on small amounts (ca. 100 pmol) of protein. Genetic engineering techniques have made it possible to construct a large number of immunoglobulins with identical binding sites for the fluorescent probe dansyllysine. These proteins differ in the heavy chain regions which are responsible for their biological effector functions and their segmental flexibility. We have analyzed a series of such constructs derived by genetic recombination between the genes coding for the mouse isotypes IgG1 and IgG2a. The results identify two regions responsible for their different degrees of segmental flexibility: the hinge region connecting the Fab and Fc portions of the antibodies, and a short stretch (residues 131-139) of sequence in the amino terminal half of the CH1 domain containing five amino acid substitutions.

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Theodore G Wensel, William P. Schneider, Vernon T Oi, and Lubert Stryel "Nanosecond Motions Of Genetically-Engineered Antibodies: Structural Elements Controlling Segmental Flexibility Defined By Time-Resolved Emission Anisotropy", Proc. SPIE 0909, Time-Resolved Laser Spectroscopy in Biochemistry, (24 June 1988); https://doi.org/10.1117/12.945374

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KEYWORDS

Proteins

Anisotropy

Fluorescence anisotropy

Polarizers

Luminescence

Polarization

Image segmentation

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