Chapter 3 Multiple Choice Questions

. Which method would you most likely choose to separate a positively-charged protein from a negatively-charged protein of roughly the same molecular weight? (Although one or more of the other choices might also work.)

Ion-exchange chromatography

Ammonium sulphate precipitation

Size-exclusion chromatography

Affinity chromatography

. For which of the following operations would size-exclusion chromatography NOT be suitable?

Separating native from denatured proteins

Separating a hexameric complex of a protein from the monomeric form

Desalting a protein

Separating positively-charged proteins from negatively-charged proteins

. Use of ammonium sulphate fractionation to separate mixtures of proteins depends on

the observation that by fixing ammonium sulphate to beads in a column, proteins that do not bind ammonium sulphate will pass through the column, but proteins that do bind ammonium sulphate will remain bound to the column.

the observation when the concentration of ammonium sulphate is raised, each protein has an individual threshold such that when the ammonium sulphate concentration reaches the threshold the protein will precipitate.

the observation that ammonium sulphate binds specifically to some proteins but not others.

the observation that positively-charged proteins bind ammonium sulphate but negatively-charged proteins do not.

. Which of the following methods of protein structure determination requires crystallization of the protein?

NMR spectroscopy

Cryo-electron microscopy

X-ray diffraction

All of the others require crystallization of the protein.

. Possible affinity tags useful for separating proteins include all but one of the following. Which one is NOT useful as an affinity tag?

His-tag

An antibody specific for the protein

In some cases, specific carbohydrates that bind to the protein

Ammonium sulphate

. Each of the following, except one, correctly states a typical difference between the results of an X-ray structure determination of a protein and an NMR structure determination of the protein. Which one is NOT a typical difference between the results of an X-ray structure determination of a protein and an NMR structure determination of the protein?

The X-ray result typically produces only one unique structure, but the NMR structure determination typically produces several models, all of which agree equally well with the experimental data.

A very high-resolution X-ray structure gives a more precise deter-mination of the coordinates of protein atoms than a typical NMR structure determination.

The X-ray result may show perturbations from the structure that actually exists free in solution because of interactions between individual molecules in a crystal, but NMR structure determinations do not show this phenomenon.

The NMR result typically produces only one unique structure, but the X-ray structure determination typically produces several models, all of which agree equally well with the experimental.

. With contemporary techniques, each of the following, except one, states a difference between cryo-electron microscopy and X-ray structure determin-ation. Which statement is incorrect?

X-ray structure determination requires crystallization of the protein (which may be difficult) whereas cryo-EM does not require crystallization of the protein.

For X-ray structure determination, it is common to travel to a synchrotron to collect data; a synchrotron is not necessary nor useful for a cryo-EM structure determination.

Cryo-electron microscopy cannot produce a structure at atomic resolution, but X-ray structure determination can.

If a protein can adopt two conformations, it may be possible to detect this and determine both conformations by cryo-EM, but to form the crystal for an X-ray structure determination may force the protein into a single conformation (even, possibly, a conformation that is not present at the highest concentration in solution).

. You wish to predict the structure of a protein of known amino-acid sequence but unknown structure. You know the structure of a related protein, such that in an optimal alignment of the sequences, 60% of the residues are the same. To do the prediction you could use

a priori prediction but not homology modelling.

either homology modelling or a priori prediction.

it would not be possible using either homology modelling or a priori prediction.

homology modelling but not a priori prediction.

. Within the past 5 years (approximately), there have been breakthrough advances in which of the following?

Neither cryo-electron microscopy nor a priori structure prediction

a priori structure prediction but not cryo-electron microscopy

Cryo-electron microscopy but not a priori structure prediction

Both cryo-electron microscopy and a priori structure prediction

. You are trying to isolate from a heterogenous mixture a protein for which the threshold for solubility is 3 M ammonium sulphate; that is, the protein is soluble if the ammonium sulphate concentration of the solution is less than 3 M but insoluble if the ammonium sulphate concentration of the solution is greater than 3M. Which of the following is correct?

You add ammonium sulphate until the concentration of ammonium sulphate is 2.5 M, then separate the precipitate from the supernatant. You keep the precipitate, and throw away the supernatant.

You add ammonium sulphate until the concentration of ammonium sulphate is 2.5 M, then separate the precipitate from the supernatant. You keep the supernatant, and throw away the precipitate.

You add ammonium sulphate until the concentration of ammonium sulphate is 0.5 M, then separate the precipitate from the supernatant. You keep the precipitate, and throw away the supernatant.

You add ammonium sulphate until the concentration of ammonium sulphate is 3.5 M, then separate the precipitate from the supernatant. You keep the supernatant, and throw away the precipitate.

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