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Biophysical Chem > test 1 > Flashcards

test 1 Flashcards

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1
Q

fluorescence polarization anisotropy

A

stable receptor-ligand complex has a large molecular mass

the complex tumbles and rotates very slowly in solution

when this complex is excited with polarized laser light, most of the fluorescing molecules emit light with a very similar polarization

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2
Q

high throughput screening (HTS) definition

A

competitive binding assay measured by fluorescence polarization anisotropy

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3
Q

high throughput screening (HTS)

A

if the compound is capable of binding more strongly to the receptor, the labelled ligand will be forced out

fluorescently labelled ligand is diffusing and rotating freely in the solution

its rotational diffusion is faster

most photons are now emitted with an arbitrary, very different polarization

by measuring this different polarization, the binding constant of each chemical compound can be determined very accurately

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4
Q

Absorption spectroscopy

A

determination of concentration of certain biomolecules

determining DNA purity

measuring oxygen saturation in blood

measuring cooperativity of hemoglobin

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5
Q

Intrinsic fluorescence spectroscopy

A

identification of the presence of fluorescing or fluorescently labelled biomolecules

Determination of metabolic activity from NADPH-fluorescence

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6
Q

fluorescence labelling

A

identification of labelled biomolecules in complex biological environments

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7
Q

FPA (fluorescence polarization anisotropy)

A

determining binding processes or other processes that can affect the rotational diffusion of labelled species

Receptor-ligand binding

molecular mass estimate

enzyme kinetics

enzyme inhibition

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8
Q

Forster resonance energy transfer (FRET)

A

determining binding processes, conformational transitions, and biomolecular distances

Receptor-ligand binding

conformation changes in DNA

determining distances during protein unfolding

membrane fusion

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9
Q

Fluorescence kinetics

A

sensing changes in the environmental polarity of biomolecules or other processes that can affect the excited state lifetime of fluorescence markers

detecting FRET or FPA via fluorescence kinetics

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10
Q

Fluorescence recovery after photobleaching

A

membrane diffusion

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11
Q

Biochemiluminescence

A

monitoring ATP concentrations

monitoring protein expression

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12
Q

Circular dichroism (CD), optical rotation dispersion

A

determination of amount of secondary structure elements in proteins

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13
Q

Light scattering

A

determination of molecular mass of biomolecules

aggregation and shape of biological objects

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14
Q

vibrational spectroscopy (infrared / Raman spectroscopy)

A

determination of secondary structure elements

label-free identification of chemical composition

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15
Q

Nuclear magnetic resonance (NMR)

A

structure determination under physiological conditions

determination of structural flexibility of biomolecules

label-free observation of biomolecular processes

comparison of NMR structure of insulin and severin with X-ray structure

flexibility of severin structure under physiological conditions

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16
Q

Electron paramagnetic resonance (EPR)

A

observation of redox reactions in photosynthesis or haemoglobin

determination of rotational diffusion using spin labels

determination of membrane structure and dynamics

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17
Q

Mass spectrometry

A

identifying biomolecules from mixtures

determining biomolecular structures from fragments

peptide sequencing

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18
Q

Fluorescence microscopy

A

imaging of labelled proteins in a whole cell context

real-time imaging of biological processes

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19
Q

Light

A

a wave of oscillating electric and magnetic fields propagating through space

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20
Q

Light field components

A

The electric and magnetic field components of electromagnetic radiation are oscillating in phase perpendicular to the propagation direction and with respect to each other

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21
Q

The smallest possible unit of light is…

A

photons

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22
Q

Photons

A

smallest possible unit of light and have particle-light and wave-light character

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23
Q

equation for energy including frequency or wavelength

A

E = hv = hc / wavelength, E = hr, v = 1 / wavelength

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24
Q

h variable

A

Planck’s constant

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25
Q

The energy of a photon is….

A

linear proportional to the frequency v and wave number

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26
Q

The wavelength of a photon is…

A

inversely proportional to the energy

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27
Q

UV is important for…

A

the characterization of proteins and DNA

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28
Q

The visible region of light is important for…

A

most fluorescence techniques

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29
Q

Infrared region is important for…

A

the identification and investigation of biomolecules based on their characteristic molecular vibrations

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30
Q

Radio-waves and microwaves are used for…

A

nuclear and magnetic resonance techniques

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31
Q

If a molecular possesses a suitable electronic structure…

A

it can absorb a photon of a specific wavelength, resulting in an electronic structure rearrangement

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32
Q

During the absorption of a photon…

A

the electronic structure of the molecules is usually transferred from an electronic ground state into an energetically higher electronic excited state

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33
Q

Only photons having __________ corresponding to _________ can be absorbed by the molecules

A

energies

the energy difference between these electronic states

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34
Q

electrons display…

A

wave-light properties

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35
Q

In phase

A

the two lines make a big squiggly line

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36
Q

out of phase

A

the two lines make a straight line

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37
Q

Molecular orbitals

A

the possible wave functions for single electrons in the presence of several nuclei

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38
Q

An orbital can only be occupies by…

A

a maximum of two electrons with opposite spins

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39
Q

If two atomic s-orbitals with opposite signs (out of phase) are combined, they form…

A

an antibonding molecular sigma orbital

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40
Q

Electrons in antibonding have…

A

higher energies

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41
Q

antibonding orbitals can lead to…

A

bond breakage

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42
Q

the energy of the newly formed molecular orbitals…

A

increases with the number of nodal planes

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43
Q

electrons from the original p-orbitals are filled into molecular orbitals, starting with…

A

the lowest energy orbitals and obeying the rule that only a maximum of two electrons are allowed per molecular orbital

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44
Q

HOMO

A

highest occupied molecular orbital

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45
Q

LUMO

A

lowest unoccupied molecular orbital

46
Q

The electronic ground state corresponds to the situation…

A

that all orbitals up to the HOMO electrons are occupied by two electrons

47
Q

The first excited state corresponds to the situation…

A

that one electron has been promoted from the HOMO to the LUMO

48
Q

The electronic ground state and the first possible electronically excited state of molecules are often referred to as…

A

S0 and S1 states

49
Q

The absorption band observed for the longest wavelength (lowest photon energy), corresponds to…

A

the absorption of photons having energies corresponding to transitions from the electronic ground state to the first possible electronically excited state (S0 - S1)

50
Q

absorption spectrum

A

describes the frequency or wavelength dependence of the probability that a molecule absorbs photons of the corresponding photon energy

51
Q

The absorption band observed for the longest wavelength corresponds to…

A

the absorption of photons having energies corresponding to transitions from the electronic ground state to the first possible electronically excited state

52
Q

The absorption band observed for the lowest photon energy corresponds to…

A

the absorption of photons having energies corresponding to transitions from the electronic ground state to the first possible electronically excited state

53
Q

The bands observed at shorter wavelengths correspond to

A

transitions in which vibrations of the nuclei in the molecules are excited in addition to the pure electronic excitation

54
Q

The bands observed at higher photon energies corresponds to

A

transitions in which vibrations of the nuclei in the molecules are excited in addition to the pure electronic excitation

55
Q

Vibrational ground state is defined as

A

v = 0

56
Q

Frank-Condon principle

A

In the first molecule, the equilibrium positions of the atoms in the excited state are very similar to those in the S0 state

No vibrations are induced by the electronic transition, and the molecule is preferentially excited into the vibrational ground state of the S1 state

In the absorption spectrum, the band corresponding to this transition dominates

57
Q

Vibrational relaxation

A

quick release of the entire vibrational excess energy in the S1 state to the surrounding

58
Q

What indicates rapid vibrational relaxation?

A

green wavy arrows

59
Q

The final result of fast vibrational relaxation is…

A

that the molecule is now in the lowest vibrational state of the electronic excited state

60
Q

The probability of fluorescence is proportional to the…

A

transition dipole moment

61
Q

The fluorescence spectrum has a …

A

mirror image like shape in comparison to the absorption spectrum

62
Q

0-0 transitions

A

the absorption and fluorescence bands with the largest and smallest wavelengths define the 0-0 transitions

63
Q

In the fluorescence spectrum, the _______ dominates

A

the vibrational band

64
Q

Absorption and fluorescence v values

A

S0, v = 0, S1, v = 0

65
Q

Stokes shift

A

the general observation that emitted photons have longer wavelengths than the absorbed photons

66
Q

If the energy gap between the s1 state and the s0 state is large

A

fluorescence dominates and the de-excitation of the moleces

67
Q

If the energy gap becomes smaller

A

nonradiative de-excitation of the s1 state often becomes dominant

68
Q

the smaller the energy gap between s1 and s0

A

the higher the chance that there is a good overlap between the vibrational ground state wave function of the s1 state with wave functions of highly excited vibrations in the electronic ground state

69
Q

Internal conversion

A

the molecule can switch from the electronic excited state into the electronic ground state without the emission of a photon

70
Q

Kasha’s rule

A

fluorescence occurs from the lowest electronically excited state in its lowest vibrational state

71
Q

Pauli rule

A

no electron in a molecule can have the exact same properties as another electron in the molecule

72
Q

An orbital can be occupied by two electrons only if…

A

they have different spins

73
Q

Triplet states

A

electronic states with two unpaired electrons of the same spin

74
Q

Triplet states are _________ compared to respective singlet states

A

lower in energy

75
Q

Intersystem crossing

A

the crossing from singlet to triplet

76
Q

Intersystem crossing is…

A

favored when a good overlap exists between vibrational wave functions of the excited S1 state, with vibrational wave functions of the T1 state

77
Q

Phosphorescence

A

radiative relaxation from the triplet state

78
Q

Phosphorescence and intersystem crossing require a…

A

spin flip

79
Q

0-0 transitions are the ______ when on a graph

A

right between the absorption last peak and the fluorescence first peak (where the mirror image begins)

80
Q

Similar to fluorescence, the phosphorescence spectrum often appears to be a ….

A

mirror image of the ground-state absorption because the vibrational modes are often very similar in all electronic states

81
Q

Jablonski diagram

A

only energy levels of the electronic states are depicted as horizontal lines

all radiative processes are depicted using straight arrows (absorption or emission of a photon)

nonradiative processes are portrayed using wavy arrows

82
Q

rate constant is proportional to

A

the probability per time unit that the corresponding process occurs

83
Q

The fluorescence quantum efficiency is

A

the number of photons emitted by a molecule as fluorescence divided by the number of photons that were previously absorbed by the moleucle

84
Q

Fluorescence quantum efficiency equation

A

number of photons emitted as fluorescence / number of absorbed photons

85
Q

quantum efficiency of any process is defined as

A

the number of quanta undergoing the process divided by the number of absorbed photons or quanta originally present in the initial state from which this process occurs

86
Q

quantum efficiency is a direct measure of

A

the probability of certain processes in a molecule

87
Q

the quantum efficiency of a process can be calculated by

A

dividing the rate constant of the process of interest by the sum of the rate constants of all processes simultaneously depopulating the initial state

88
Q

to calculate the phosphorescence quantum yield

A

you have to multiply the quantum yield for s1 - t1 intersystem crossing by the probability for radiative emission from the triplet state

89
Q

the sum of all radiative and nonradiative processes…

A

must be one

90
Q

time constant

A

inverse of its corresponding rate constant

91
Q

the absorption of a compound for a certain wavelength can be evaluated by..

A

a comparison between the light intensity measures after the sample cell and the light intensity measured after the reference cell

92
Q

wavelength-dependent transmission equation

A

T (wavelength) = light intensity after sample cell / light intensity after reference cell

93
Q

the reduction of the light intensity is…

A

proportional to the concentration of the absorbing compound and the length of the sample cell

94
Q

Beer-Lambert Law

A

log (light intensity reference cell / light intensity sample cell) = the molar absorption * concentration * total optical path length of the light through the sample cell

95
Q

The absorption of a compound at a given compound is defined as

A

A = ECL

96
Q

the molar absorption coefficient is proportional to

A

the corresponding electronic transition dipole moment

97
Q

Transition-dipole moment describes

A

the magnitude of absorption

98
Q

The Franck-Condon integral descibres

A

which wavelength will result in absorption maxima

99
Q

the fluorescence intensity is proportional to

A

the concentration of the compound

100
Q

proteins and peptides show ___________

A

unspecific absorptions at wavelengths below 240nm

101
Q

which amino acids have distinctive absorbance and fluorescence properties

A

tryptophan, tyrosine, and phenylalanine

102
Q

Redshifts

A

shifts of absorption or fluorescence bands towards longer wavelengths (blueshifts are the opposite)

103
Q

DNA and RNA both have _______ that show ______

A

conjugates pi electron systems

significant absorption around 260nm

104
Q

pure DNA value is

A

1.8

105
Q

pure RNA value is

A

2.0

106
Q

when you want to run a blank sample on the spectrophotometer, you should use

A

whatever your sample is dissolved into

107
Q

easy way to remember Beer’s law

A

A = ECL

108
Q

haem is a cofactor for

A

protein hemogloblin, which is responsible for the transportaion of oxygen in red blood cells

109
Q

the absorption change between _____________ is used for _______

A

hemoglobin and oxyhemoglobin

determine the oxygen saturation in a patient’s blood

110
Q

flavins

A

an important class of cofactors that dominate the optical signals from cells

Biophysical Chem (2 decks)

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