1. BCHM - Structural Biology
Question Information
Q-Bank
MCAT
Sim
Non-Sim
Subject
Biochemistry
Foundation
BCHM - Structural Biology
Validity
5 years
Author(s)
Reyes, V. M.
Reviewer(s)
Editor(s)
Passage Media
Media ID(s)
Passage
If we had x-ray vision, or, alternatively, an x-ray microscope, we would be able to
actually “see” molecules, including protein molecules, changing slightly in shape
(much like living, breathing entities, inhaling and exhaling!) and undergoing conformational changes as they move around. But such a scenario will never happen any time soon, so, for biomolecules at least, we rely on x-ray crystallography (XRC).
As its name implies, to do XRC we need an x-ray source that is coherent (emitting x-rays that all have the same wavelength), and a crystalline form of the molecule whose
structure we want to “see”. Many techniques have been developed for the formation
of protein crystals, and the literature on protein crystallization is quite large.

2. Figure 1 Tetragonal crystals of enzyme (protein), lysozyme.
Figure 1 shows crystals of protein lysozyme, an enzyme that catalyzes the breakdown
by hydrolysis of certain polysaccharides. Protein crystals produced by current
technology in XRC are barely visible to the eye and are often less than 1 mm. in cross-section, so a desk microscope is often required for its routine close examination during growth.
A “good” protein crystal must be molecularly ordered enough within the crystal lattice
to be able to diffract an x-ray beam at acceptably high angles. The higher the angles
a protein crystal diffracts an incident x-ray beam to, the higher the resolution - i.e., the
finer the detail – the “image” of the protein in the crystal will be in the final result
(the model).
Figure 2 shows a (very) simplified set-up for an XRC experiment. The x-ray source may be an in-house x-ray generator, or a government-run synchrotron facility (better). The protein crystal itself is within a droplet of crystallization solution on a small wire loop mounted on a device called a goniometer. The goniometer allows the rotation of the crystal at any angle on any axis: this is to be able record as many diffraction spots on the detector as possible. These diffraction spots are the crucial information one needs to be able to “reconstruct” the atomic structure of the molecule making up the crystal that gave rise to the diffraction spots themselves.

3. How is the image reconstruction done
How is the image reconstruction done? The theory behind it is a bit complicated and
mathematical but in a nutshell, one iteratively employs forward and reverse Fourier
transformations on the diffraction spots, using their intensities and relative positions
in the calculations. Of course these days all of these calculations are performed by high-
end computers, and the crystallographer nowadays need only know how to run soft-
ware to do these tricks.
Figure 2
After careful statistical analysis of the results to make sure that the data fits the model,
the result is an image of the protein in the form of a more or less continuous
electron cloud representing the amino acids making up the protein chain folded in
three-dimension. For example, where there is a phenylalanine along the
polypeptide chain, one “sees” an electron cloud in the shape of a phenylalanine R-
group at that site. The rest of the work involves modeling the atomic positions by
assigning Cartesian (x,y,z) coordinates to each (or almost) of them.

4. A. Lehninger, Principles of Biochemistry
Passage References
PMID/Book
Title of Publication or Book
A. Lehninger, Principles of Biochemistry
(N/A)
en.wikipedia.com, youtube.com
(N/A)
Author’s own lecture notes.
Question Attributes #1
Topic Blueprint
Macromolecular X-Ray Crystallography
Competency
MCAT: BS-3: Scientific research methodology
To understand the basic technology involved in protein
(macromolecular) x-ray crystallography.
Objective
Media ID(s)
Question ID
Question Stem #1
Which of the following statements is false about protein structure determination by x-ray crystallography?
Answer Choices #1
X-ray crystallography may also be used to solve the 3D structures
of lipids, carbohydrates, nucleic acids (DNA and RNA) and viruses. A)
The 3D structures solved through x-ray crystallography should always
be taken with a grain of salt, since biomolecules in the cell are not in
a crystalline environment but in an aqueous one. B)

5. Answer Choices #1 (cont’d.)
Structures of membrane proteins are in general the most difficult to
solve using XRC since they are difficult to crystallize due to their
hydrophobicity. C)
Crystals that diffract very well implies that the protein molecules
within the crystal lattice are well-ordered. D)
Correct: B)
Explanation #1
The correct answer is choice B. The question requires the student to select the
false statement about protein structure determination by x-ray crystallography,
and statement B is false. It has been determined that the water content in protein
crystals is the same as that in the cytoplasm. That being so, it is no surprise that
protein 3D structures determined by XRC are the true structures that the protein
assumes in the cell. Protein crystal x-ray structures are accurate, and only in very
rare instances do they differ from solution structures, such as those determined by
protein NMR.
(Choice A) This statement is true: if you can crystallize it, you can use XRC to
determine its structure, no matter if it is lipid, carbohydrate, nucleic acid (DNA or
RNA) or even viruses.
(Choice C) This statement is also true. Since membrane proteins have to interact with
the lipid bilayer (which is hydrophobic), they also need to be hydrophobic. Hydrophobic
proteins tend to repel each other and resist being in the crystal lattice. Special additives
need to be added to the crystallization solution to achieve stability within the crystal.
(Choice D) This statement is true. Crystals that do not diffract very well (i.e., all
diffraction spots are low-angle) have a high degree of mosaicity, meaning that
the molecules within the crystal lattice are disordered.
Educational objective: To understand the basic technology involved in protein
(macromolecular) x-ray crystallography.

6. 0000000 0000000 References #1 PMID/Book Title of Publication or Book
Verifications #1
Yes / No
The question is at the Application or higher cognitive level.
Yes / No
The question is based on a realistic clinical scenario.
Yes / No
The question has at least one close distracter, and other options have
educational value.
Yes / No
The question is appropriate to the entry level of nursing practice.
Yes / No
The explanation is short and concise, yet thorough.
Yes / No
The question has an appropriate table/flow chart/illustration.