Question

•   Describe why Bragg’s law is nλ = 2dsinΘ. Diagram will be given.

•   Describe how electrons play a role in NMR.

•   Describe, simply, how NMR allows us to get a 3-D structure of a protein.

•   Describe the basic process of cryo-EM

Answer 1

1, Bragg law, in physics, the relation between the spacing of atomic planes in crystals and the angles of incidence at which these planes produce the most intense reflections of electromagnetic radiations, such as X rays and gamma rays, and particle waves, such as those associated with electrons and neutrons. For maximum intensity of reflected wave trains, they must stay in phase to produce constructive interference, in which corresponding points of a wave (e.g., its crests or troughs) arrive at a point simultaneously. The Bragg law was first formulated by Lawrence Bragg, an English physicist. Bagg's Law refers to the simple equation:

nλ = 2d sinΘ

derived by the English physicists Sir W.H. Bragg and his son Sir W.L. Bragg in 1913 to explain why the cleavage faces of crystals appear to reflect X-ray beams at certain angles of incidence (Θ, λ). The variable d is the distance between atomic layers in a crystal, and the variable lambda is the wavelength of the incident X-ray beam (see applet); n is an integer.

2. The power of NMR is based on the concept of nuclear shielding, which allows for structural assignments. Every atom is surrounded by electrons, which orbit the nucleus. Charged particles moving in a loop will create a magnetic field which is felt by the nucleus. Therefore the local electronic environment surrounding the nucleus will slightly change the magnetic field experienced by the nucleus, which in turn will cause slight changes in the energy levels! This is known as shielding. Nuclei that experinece differnet magnetic fields due to the local electronic interactions are known as inequivalent nuclei. The change in the energy levels requires a different frequency to excite the spin flip, which as will be seen below, creates a new peak in the NMR spectrum. The shielding allows for structural determination of molecules.

3. Measures the energy levels of magnetic atoms, i.e. atoms with odd electron numbers: 1H, 13C, 15N, 19F, 31P Energy levels of an atom are influenced by the local environment (chemical shifts) Via covalent bonds , Through space, max. 5A apart: Nuclear Overhauser Effect (NOE) , NMR can identify atoms that are close together, also those that are close in space but not linked by direct covalent bonds, Chemical shifts can define secondary structures, NMR spectra yield a set of peaks that correspond to the interactions between pairs of atoms  From these, one can calculate the protein structure

4. Cryo-electron microscopy (cryoEM) is an ensemble of techniques allowing the observation of biological specimens in their native environment at cryogenic temperatures in EM (-180°C for liquid nitrogen stages, -269°C for He).Cryo-electron microscopy (cryo-EM) is increasingly becoming a mainstream technology for studying the architecture of cells, viruses and protein assemblies at molecular resolution.Over the past decade, the phrase “cryo-electron microscopy”, often abbreviated as “cryo-EM”, has evolved to encompass a broad range of experimental methods. At the core, each of these is based upon the principle of imaging radiation-sensitive specimens in a transmission electron microscope under cryogenic conditions. In biology, applications of cryo-EM now span a wide spectrum, ranging from imaging intact tissue sections and plunge-frozen cells to individual bacteria, viruses and protein molecules. Cryo-electron tomography, single-particle cryo-electron microscopy, and electron crystallography are all sub-disciplines of cryo-EM that have been used successfully to analyze biological structures in different contexts. These methods have been used singly as well as in hybrid approaches, where the information from electron microscopy is combined