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Picosecond fluorescence spectroscopy of a single-chain class I major histocompatibility complex encoded protein in its peptide loaded and unloaded states.

Abstract：

:The tryptophan fluorescence properties of two different peptide complexes of the single-chain H-2Kd (SC-Kd) were studied by means of the single-photon counting technique. The latter enables time-resolved measurements of fluorescence intensity and anisotropy decay parameters relevant to structural and dynamic properties of proteins. While the isolated SC-Kd molecules in their 'original' purified form represent the unloaded state, i.e., containing endogenous low-affinity peptides, the loaded SC-Kd protein is obtained by introducing well-defined high-affinity peptides that replace the low-affinity ones. These two SC-Kd forms were found to exhibit different time-resolved tryptophan emission patterns; the unloaded complexes show a slightly faster fluorescence intensity decay rate than the loaded one. Three well-resolved time domains were distinguished in the anisotropy decay course of both forms: a short one in the picosecond range, an intermediate one of several nanoseconds, and a long one spanning several dozens to hundreds of nanoseconds. They are assigned to superposition contributions of (short- and long-distance) non-radiative energy transfer processes, to motions of the tryptophans, and to rotation of the whole protein globule. In the loaded SC-Kds, the first two processes were found to be attenuated. It is therefore suggested that upon binding of high-affinity peptides, the SC-Kd structure becomes more compact and certain tryptophans become less accessible to quenchers. The faster anisotropy decay observed in the unloaded form reflects both an enhancement in the energy-transfer between the tryptophans and an acceleration of their motions.(ABSTRACT TRUNCATED AT 250 WORDS)

journal_name

Immunol Lett

journal_title

Immunology letters

authors

Dittes K,Gakamsky DM,Haran G,Haas E,Ojcius DM,Kourilsky P,Pecht I

doi

10.1016/0165-2478(94)90183-x

subject

Has Abstract

pub_date

1994-05-01 00:00:00

pages

125-32

issue

eissn

0165-2478

issn

1879-0542

pii

0165-2478(94)90183-X

journal_volume

pub_type

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Picosecond fluorescence spectroscopy of a single-chain class I major histocompatibility complex encoded protein in its peptide loaded and unloaded states.