Phasing

Question 1

Dense Modification techniques

Answer 1

Solvent flattening
Solvent flipping
NCS averaging
Averaging between crystal forms
Histogram matching

Question 2

Molecular Replacement “good” model demands

Answer 2

Similarity between query and target
Represent enough space
Placed in correct orientation
No water, ions

Question 3

lack-of-closure

Answer 3

The distance between real phase and calculated.

Question 4

m

Answer 4

Figure of merit for phase.
“is the vector from the
centre of the circle to C. It is a measure of how
well determined the phase angle is.”

Question 5

Isomorphous replacement gives?

Answer 5

minimal changes in protein

Question 6

SIR

Answer 6

one derivative – phase ambiguity (two phases)

Question 7

MIR

Answer 7

multiple derivatives – phase ambiguity resolved

Question 8

Heavy atom sub-structure solved how?

Answer 8

using difference Patterson map
delta(Fiso) = | Fph - Fp | ≈ Fh

Question 9

Friedels law

Answer 9

F(hkl) = F(-h-k-l)
alpha(hkl) = -alpha(-h-k-l)

Question 10

Anomalous scattering

Answer 10

Difference between F(hkl) and F(-h-k-l) can be used for phasing

If the incident beam has a wavelength close to an absorption edge of an atom in the crystal, Friedel’s law breaks down and is no longer true.

Question 11

Solvent flattening

Answer 11

Identify boundary between protein and solvent (mask = Wang)
Set solvent density to constant value (k=0)
Calculate phases (adm) and combine with aexpt
Calculate new map using Fobs and acomb
Repeat until convergence

Question 12

Solvent flipping

Answer 12

k < 0
Often produces good results unless solvent content very high
Similar to adding negative noise to an image in order to strengthen the
signal/noise ratio
Reduces bias

Question 13

NCS averaging

Answer 13

Non-crystallographic symmetry (NCS) averaging
Crystals and lattices can only have 2-, 3-, 4-, and 6-fold rotational symmetry
For each point in P1:
Find the NCS equivalent points in P2, P3, … , PN
Calculate the average density rave = (rP1 + rP2 + rP3 + rPN)/N
Assign the average density to each of the equivalent points
*Flatten (or flip) the solvent region
*Back-transform to get Fdm, adm
*Combine adm with aexpt to get acomb
*Use acomb with Fobs to calculate new map, repeat until convergence

Question 14

Averaging between different crystal forms

Answer 14

Protein crystallizes in more than one crystal form, averaging between can give better phase.
Construct averaged (and solvent flattened) maps for each crystal form
Back-transform each to get Fdm, adm for each crystal form
Combine adm with aexpt to get acomb
Use acomb with Fobs to calculate new maps, repeat until convergence

Question 15

Histogram matching

Answer 15

Modification of the electron densities of an initial map to match an ideal histogram

Question 16

Origin shifts

Answer 16

Substructures with different origins give identical maps

Question 17

Both enantiomers

Answer 17

One hand is correct and will give at least decent estimates of the phases, and a ~decent map. Other horrible phases :)

Question 18

Best fourier

Answer 18

Phase angle at the centroid.
Fbest = mFobsexp(i*abest)
For each reflection, we use observed (measured) structure factor amplitude (F obs) and the phase angle at the centroid of the distribution (α best), and we also weight each Fourier term with the figure-of-merit (m) for the phase angle.

Question 19

SAD

Answer 19

Single-wavelength anomalous dispersion, using data at Df” peak.

Question 20

MAD

Answer 20

Multiple-wavelength anomalous dispersion, data sets measured with lambda, and on each side of, the Df” peak.