Construction and Performance of a Fluorescence Polarization Spectrophotometer

Weber, Gregorio; Bablouzian, B.

doi:10.1016/s0021-9258(18)96575-0

Cited by 105 publications

(27 citation statements)

References 9 publications

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“…Corrected fluorescence spectra were obtained with a Turner 210 spectrofluorimeter using cells of 1-cm optical path, the optical density of the solution being less than 0.1 at the excitation wavelength. Fluorescence polarization spectra1 were obtained with an instrument of the type described by Weber and Bablouzian (1966), built by Dr. M. Shinitzky, the 1 The fluorescence polarization p is defined as (/|| -/j_)/(/y + /_l ), where /¡¡ and /± are the fluorescence intensities observed through a polarizer oriented parallel and perpendicular, respectively, to the direction of polarization of the exciting light.…”

Section: Methodsmentioning

confidence: 99%

Fast relaxation processes in a protein revealed by the decay kinetics of tryptophan fluorescence

Grinvald¹,

Steinberg²

1974

Biochemistry

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AHSTRAC r: The fluorescence decay of chicken pepsinogen is not monoexponential throughout the emission spectrum. For light emitted a t the long wavelength region of the fluorescence spectrum, the decay can be described by two exponential terms, one of them exhibiting a negative amplitude. l'his behavior shows that in this region of the spectrum the fluorescence builds up before it decays, indicating that the electronically excited species involved has been created during the fluorescence lifetime. For a variety of reasons it seems very unlikely that this emitting species as formed by energy transfer from other entities in the protein. The buildup of the fluorescence at the red edge of the spectrum prior to emission thus reflects a genuine relaxation process i n the protein molecule in the nanosecond time scale. the exc,i:ed chromophore shifting its emission spectrum to the red in the course of the relaxation process. The reaction in-T f i e fluorescence emitted by molecules in condensed phases is invariably shifted to longer wavelengths relative to the absorption spectrum. The larger the interaction between the excited chroniophore with its environment the greater the shift of the fluorescence band to the red. This behavior of fluorescence spectra is due to relaxation processes which the excited molecule undergoes prior to the emission process, the rate of relaxation being usuallj. much faster than the rate of light emission. The processes which take place in the excited state may be of a variety of kinds: dissipation of vibrational energy or change of conformation of the electronicall4 excited molecules, interactions with surrounding solvent molecules, association or dissociation reactions in the excited state, or processes of fast energy transfer from one chrornophoric group to another in the system studied. Part of the energy of the excited molecules is lost in these processes and the photon emitted subsequently is left N i t h less energ) t o carry; the emitted light is thus of longer wavelerig t h .I n all cases in which the reactions in the excited state are much faster than the rate of light emission, the fluorescence collected at the various wavelengths of the spectrum deca) s ni ono t on I) us1 y with ti iri e. IJ nder certain circumstances son1 e of t h e relaxation processes are not fast compared t o the lifetime of the excited state. Such a situation is disclosed by buildup of the fluorescence intensity prior to the decay at some region of the emission spectrum. Examples of this kind were reported when the electronically excited molecule undergoes association reactions (Speed and Selinger. 1969) volved in the relaxation may be a nonspecific orientation of various groups around the excited chromophore or a forniation of a more specific excited state complex, i.e.. an exciplex. This nanosecond relaxation process is Conformation dependent and disappears upon denaturation. Similarly, chicken pepsin a t neutral pH fails to show it. This sce1m to be the first case in which a relaxation process in the iianos...

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Section: Methodsmentioning

confidence: 99%

Fast relaxation processes in a protein revealed by the decay kinetics of tryptophan fluorescence

Grinvald¹,

Steinberg²

1974

Biochemistry

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“…The 1 Abbreviations used are: ANS, l-anilino-8-naphthalenesulfonate; DNS, l-dimethylamino-5-naphthalenesulfonate. emitted light passed through liquid filters of 2 m NaNo2 and Corning glass 3-72; (ii) with the instrument designed by Weber and Bablouzian (1966) for measurements in the ultraviolet region. This instrument was used to record fluorescence polarization spectra between 250 and 320 nm.…”

Section: Methodsmentioning

confidence: 99%

Fluorescence studies of Aplysia and sperm whale apomyoglobins

Anderson¹,

Brunori²,

Weber³

1970

Biochemistry

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“…built by him (Weber and Young, 1964). The polarization spectra were obtained with the instrument described by Weber and Bablouzian (1966). The cuvet compartments of both instruments were maintained at constant temperature by means of a circulating water bath.…”

Section: Methodsmentioning

confidence: 99%

Effects of Temperature and Activating Cations on the Fluorescence of Pyruvate Kinase^*

Ch¹

1967

Biochemistry

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Construction and Performance of a Fluorescence Polarization Spectrophotometer

Cited by 105 publications

References 9 publications

Fast relaxation processes in a protein revealed by the decay kinetics of tryptophan fluorescence

Fast relaxation processes in a protein revealed by the decay kinetics of tryptophan fluorescence

Fluorescence studies of Aplysia and sperm whale apomyoglobins

Effects of Temperature and Activating Cations on the Fluorescence of Pyruvate Kinase^*

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Construction and Performance of a Fluorescence Polarization Spectrophotometer

Cited by 105 publications

References 9 publications

Fast relaxation processes in a protein revealed by the decay kinetics of tryptophan fluorescence

Fast relaxation processes in a protein revealed by the decay kinetics of tryptophan fluorescence

Fluorescence studies of Aplysia and sperm whale apomyoglobins

Effects of Temperature and Activating Cations on the Fluorescence of Pyruvate Kinase*

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Effects of Temperature and Activating Cations on the Fluorescence of Pyruvate Kinase^*