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  Dynamics of molecules involved in antigen presentation: effects of fixation

Barisas, B. G., Wade, W. F., Jovin, T. M., Arndt-Jovin, D. J., & Roess, D. A. (1999). Dynamics of molecules involved in antigen presentation: effects of fixation. Molecular Immunology, 36, 701-708.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0012-FA2A-7 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-0027-E825-3
Genre: Journal Article

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 Creators:
Barisas, B. G., Author
Wade, W. F., Author
Jovin, T. M.1, Author              
Arndt-Jovin, D. J.1, Author              
Roess, D. A., Author
Affiliations:
1Department of Molecular Biology, MPI for biophysical chemistry, Max Planck Society, ou_578628              

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Free keywords: Class-ii molecules; T cells; Rotational diffusion; Mast cells; Mhc; Phosphorescence; Recognition; Anisotropy; Receptors; Surface
 Abstract: Antigen presentation by MHC class II molecules can be enhanced by paraformaldehyde fixation of antigen-presenting cells prior to assay. This treatment might be expected to aggregate membrane proteins and thus stabilize and strengthen transient protein-protein interactions involved in intercellular cooperation. Lateral and rotational dynamics of the MHC class II antigen I-A(d) on A20 cells fixed with various concentrations of paraformaldehyde were examined by fluorescence photobleaching recovery and time-resolved phosphorescence anisotropy, respectively. Probes were tetramethylrhodamine and erythrosin conjugates of MKD6 Fab fragments. Increasing concentrations of paraformaldehyde led to a progressive increase in the limiting anisotropy of I-A(d) at 4 degrees C from the value of 0.042 for untreated cells, indicative of large aggregate formation, while leaving the rotational correlation time of 29 mu s unchanged, a measure of the unperturbed molecule. On the other hand, the translational diffusion constants decreased from similar to 2 x 10(-10) cm(2) s(-1), while me fractional recovery remained unchanged at about 40-50%. Taken together, these results suggest that fixation crosslinks class II molecules to each other or to other membrane proteins into structures large enough (> 500,000 kDa) to diffuse translationally with perceptibly size-dependent rates. The fixation effects on both class II rotation and lateral diffusion were half-maximal at paraformaldehyde concentrations of similar to 0.2%, Possible relations between the biological effector functions of class II and the physical sizes of fixation-induced aggregates are discussed. (C) 1999 Elsevier Science Ltd. All rights reserved.

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Language(s): eng - English
 Dates: 2005-07-081999
 Publication Status: Published in print
 Pages: -
 Publishing info: -
 Table of Contents: -
 Rev. Method: Peer
 Identifiers: eDoc: 223142
Other: 11464
 Degree: -

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Title: Molecular Immunology
Source Genre: Journal
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Pages: - Volume / Issue: 36 Sequence Number: - Start / End Page: 701 - 708 Identifier: -