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Analysis of time-resolved fluorescence anisotropy in lipid-protein systems

II. Application to tryptophan fluorescence of bacteriophage M13 coat protein incorporated in phospholipid bilayers

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Abstract

The subnanosecond fluorescence and motional dynamics of the tryptophan residue in the bacteriophage M13 coat protein incorporated within pure dioleoylphosphatidylcholine (DOPC) as well as dioleoylphosphatidylcholine/dioleoylphosphatidylglycerol (DOPC/DOPG) and dimyristoylphosphatidylcholine/dimyristoylphosphatidylglycerol (DMPC/DMPG) bilayers (80/20 w/w) with various L/P ratios have been investigated. The fluorescence decay is decomposed into four components with lifetimes of about 0.5, 2.0, 4.5 and 10.0 ns, respectively. In pure DOPC and DOPC/DOPG lipid bilayers, above the phase transition temperature, the rotational diffusion of the protein molecules contributes to the depolarization and the anisotropy of tryptophan is fitted to a dual exponential function. The longer correlation time, describing the rotational diffusion of the whole protein, shortens with increasing temperature and decreasing protein aggregation number. In DMPC/DMPG lipid bilayers, below the phase transition, the rotational diffusion of the protein is slowed down such that the subnanosecond anisotropy decay of tryptophan in this system reflects only the segmental motion of the tryptophan residue. Because of a heterogeneous microenvironment, the anisotropy decay must be described by three exponentials with a constant term, containing a negative coefficient and a negative decay time constant. From such a decay, the tryptophan residue within the aggregate undergoes a more restricted motion than the one exposed to the lipids. At 20°C, the order parameter of the transition moment of the isolated tryptophan is about 0.9 and that for the exposed one is about 0.5.

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Abbreviations

DOPC:

1,2-dioleoyl-sn-glycero-3-phosphocholine

DOPG:

1,2-dioleoyl-sn-glycero-3-phosphoglycerol

DMPC:

1,2-dimyristoyl-sn-glycero-3-phosphocholine

DMPG:

1,2-dimyristoylsn-glycero-3-phosphoglycerol

L/P ratio:

phospholipid to coat protein molar ratio

<τ>:

average fluorescence lifetime

r (0):

initial anisotropy

r (∝):

residual anisotropy

S :

order parameter of the transition moment of the tryptophan residue

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Authors and Affiliations

  1. Department of Molecular Physics, Agricultural University, P.O. Box 8128, 6700 ET, Wageningen, The Netherlands

    K. Peng, A. van Hoek, C. J. A. M. Wolfs & M. A. Hemminga

  2. Department of Biochemistry, Agricultural University, P.O. Box 8128, 6700 ET, Wageningen, The Netherlands

    A. J. W. G. Visser

Authors
  1. K. Peng
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  2. A. J. W. G. Visser
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  3. A. van Hoek
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  4. C. J. A. M. Wolfs
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  5. M. A. Hemminga
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On leave of Shanghai Medical Equipment Research Institute, 77 Jiang Ning Rd. Shanghai, People's Republic of China

Offprint requests to: M. A. Hemminga

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Peng, K., Visser, A.J.W.G., van Hoek, A. et al. Analysis of time-resolved fluorescence anisotropy in lipid-protein systems. Eur Biophys J 18, 285–293 (1990). https://doi.org/10.1007/BF00188041

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  • DOI: https://doi.org/10.1007/BF00188041

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