Transcript Online Counseling Resource YCMOU ELearning Drive…

Online Counseling Resource
YCMOU ELearning Drive…
School of Architecture, Science and Technology
Yashwantrao Chavan Maharashtra
Open University, Nashik – 422222, India
OC-SBI074-CP1-02
Introduction
Programmes and Courses
 SEP –SBI074-CP1_U01
School of Science and Technology, Online Counseling Resource…
Credits
 Academic Inputs by
 Sonali Alkari
 MSc (Botany), P.G. D.C. Bio-Informatics
[email protected]
School of Science and Technology, Online Counseling Resource…
How to Use This Resource

Counselor at each study center should use this presentation to deliver
lecture of 40-60 minutes during Face-To-Face counseling.

Discussion about students difficulties or tutorial with assignments should
follow the lecture for about 40-60 minutes.

Handouts (with 6 slides on each A4 size page) of this presentation should
be provided to each student.

Each student should discuss on the discussion forum all the terms which
could not be understood. This will improve his writing skills and enhance
knowledge level about topics, which shall be immensely useful for end
exam.

Appear several times, for all the Self-Tests, available for this course.

Student can use handouts for last minutes preparation just before end
exam.
© 2007, YCMOU. All Rights Reserved.
4
School of Science and Technology, Online Counseling Resource…
Learning Objectives
 After studying this module, you should be able
to:
 Explain Single-crystal X-ray Diffraction
 Describe Fundamental Principles of Singlecrystal X-ray Diffraction
 Explain working of Single-crystal X-ray
Diffraction Instrumentation
 State Strengths and Limitations of Singlecrystal X-ray Diffraction
© 2007, YCMOU. All Rights Reserved.
5
School of Science and Technology, Online Counseling Resource…
Introduction:1
 X-ray scattering techniques are a family of nondestructive
analytical
techniques
which
reveal
information about the crystallographic structure,
chemical composition, and physical properties of
materials and thin films.
 These techniques are based on observing the scattered
intensity of an x-ray beam hitting a sample as a function
of incident and scattered angle, polarization, and
wavelength or energy.
 To
characterize the crystallographic structure of
crystalline material and powdered solid samples
different diffraction techniques are used
School of Science and Technology, Online Counseling Resource…
Introduction:2
 Single-crystal X-ray diffraction is a technique
used to solve the complete structure of
crystalline materials, ranging from simple
inorganic solids to complex macromolecules,
such as proteins.
 Powder diffraction (XRD) is a technique use to
characterize the crystallographic structure,
crystallite size (grain size), and preferred
orientation in polycrystalline or powdered
solid samples.
School of Science and Technology, Online Counseling Resource…
X-ray Scattering Techniques:1
 X-ray scattering techniques are a family of nondestructive analytical techniques.
 X-ray scattering techniques which reveal
information
about
the
crystallographic
structure, chemical composition, and physical
properties of materials and thin films.
 These techniques are based on observing the
scattered intensity of an x-ray beam.
 The X-ray beam hits the sample as a function of
incident and scattered angle, polarization, and
wavelength or energy.
School of Science and Technology, Online Counseling Resource…
X-ray Scattering Techniques:2
 X-ray diffraction techniques are based on the
elastic scattering of x-rays from structures that
have long range order.
 The
most
comprehensive
description
of
scattering from crystals is given by the
dynamical theory of diffraction.
 Max von Laue, in 1912, discovered that
crystalline substances act as three-dimensional
diffraction gratings for X-ray wavelengths
similar to the spacing of planes in a crystal
lattice.
School of Science and Technology, Online Counseling Resource…
X-ray Diffraction Pattern
 This
is
an
X-ray
diffraction
pattern
formed when X-rays
are
focused
on
a
crystalline material.
 Each dot, called a
reflection, forms from
the
coherent
interference
of
scattered
X-rays
passing through the
crystal.
School of Science and Technology, Online Counseling Resource…
Single-crystal X-ray Diffraction-1
 Single-crystal X-ray Diffraction is a nondestructive analytical technique.
 Single crystal X-ray diffraction provides
detailed information about the internal lattice
of crystalline substances.
 Single crystal X-ray diffraction provide details
about
unit cell dimensions, bond-lengths,
bond-angles, and details of site-ordering.
 Single crystal X-ray diffraction
is directly
related is single-crystal refinement.
 In
single-crystal
refinement,
the
data
generated
from
the
X-ray
analysis
is
interpreted and refined to obtain the crystal
structure.
School of Science and Technology, Online Counseling Resource…
Single Crystal X-ray Diffraction-2
 The most common experimental method of obtaining a
detailed structure of a molecule is single crystal X-ray
diffraction (SXRD) .
 Single crystal X-ray diffraction (SXRD)
allows
resolution of individual atoms.
 single crystal X-ray diffraction (SXRD) performed by
analyzing the pattern of X-rays diffracted by an ordered
array of many identical molecules (single crystal).
 Many pure compounds, from small molecules to
organometallic complexes, proteins, and polymers,
solidify into crystals under the proper conditions.
School of Science and Technology, Online Counseling Resource…
Single Crystal X-ray Diffraction-3
 When solidifying into the crystalline state, these
individual molecules typically adapt one of the few
possible 3D orientations.
 When a monochromatic X-ray beam is passed through
a single crystal, the radiation interacts with the
electrons in the atoms, resulting in scattering of the
radiation to produce a unique image pattern.
 Multiple images are recorded, with an area X-ray
detector, as the crystal is rotated in the X-ray beam.
 Computationally intensive analysis of a set images
results in a solution for the 3D structure of the
molecule.
School of Science and Technology, Online Counseling Resource…
Principles of X-ray Diffraction:1
 X-ray diffraction is based on constructive
interference of monochromatic X-rays and a
crystalline sample.
 These X-rays are generated by a cathode ray
tube, filtered to produce monochromatic
radiation, collimated to concentrate, and
directed toward the sample.
 The interaction of the incident rays with the
sample produces constructive interference
(and a diffracted ray) when conditions satisfy
Bragg's Law (nλ=2d sinθ).
School of Science and Technology, Online Counseling Resource…
Principles of X-ray Diffraction:2
 This law relates the wavelength of
electromagnetic
radiation
to
the
diffraction angle and the lattice spacing
in a crystalline sample.
 These
diffracted
X-rays
are
then
detected, processed and counted.
 By changing the geometry of the
incident rays, the orientation of the
centered crystal and the detector, all
possible diffraction directions of the
lattice should be attained.
School of Science and Technology, Online Counseling Resource…
Principles of X-ray Diffraction:3
 All diffraction methods are based on
generation of X-rays in an X-ray tube.
These X-rays are directed at the sample,
and the diffracted rays are collected.
 A key component of all diffraction is the
angle
between
the
incident
and
diffracted rays.
 Powder and single-crystal diffraction
vary in instrumentation.
School of Science and Technology, Online Counseling Resource…
Instrumentation of X-ray Diffraction
 X-ray diffractometers consist of three basic
elements, an X-ray tube, a sample holder, and
an X-ray detector.
 X-rays are generated in a cathode ray tube by
heating a filament to produce electrons,
accelerating the electrons toward a target by
applying a voltage, and impact of the
electrons with the target material.
 When electrons have sufficient energy to
dislodge inner shell electrons of the target
material, characteristic X-ray spectra are
produced.
 X-rays must be produced using a synchotron,
which emits a much stronger beam.
School of Science and Technology, Online Counseling Resource…
How Does It Work?
 These spectra consist of several
components, the most common being
Kα and Kβ.
 Kα consists, in part, of Kα1 and Kα2.
 Kα1 has a slightly shorter wavelength
and twice the intensity as Kα2.
 In case of Molybdenum, which is a
common target material for singlecrystal diffraction,
Kα radiation = 0.7107Å.
School of Science and Technology, Online Counseling Resource…
How Does It Work - 1
 For diffraction, monochromatic Xrays
are
needed
which
are
produced by filtering, by foils or
crystal.
 Kα1and Kα2 are sufficiently close in
wavelength such that a weighted
average of the two is used.
 The specific wavelengths are
characteristic
of
the
target
material.
School of Science and Technology, Online Counseling Resource…
How Does It Work - 3
 When the geometry of the incident X-rays
impinging the sample satisfies the Bragg
Equation, constructive interference occurs.
 A detector records and processes this X-ray
signal and converts the signal to a count rate
which is then output to a device such as a
printer or computer monitor.
 Modern single-crystal diffractometers use CCD
(charge-coupled
device)
technology
to
transform the X-ray photons into an electrical
signal which are then sent to a computer for
processing.
School of Science and Technology, Online Counseling Resource…
How Does It Work - 4
 Single-crystal
diffractometers use either
3- or 4-circle goniometers.
 These circles refer to the
four angles (2θ, χ, φ, and
Ω)
that
define
the
relationship between the
crystal lattice, the incident
ray and detector.
 Samples are mounted on
thin glass fibers which are
attached to brass pins and
mounted onto goniometer
heads.
 Adjustment of the X, Y
and
Z
orthogonal
directions
allows
centering of the crystal
within the X-ray beam.
School of Science and Technology, Online Counseling Resource…
How Does It Work - 5
 X-rays leave the collimator and are directed at
the crystal.
 Rays are either transmitted through the
crystal, reflected off the surface, or diffracted
by the crystal lattice.
 A beam stop is located directly opposite the
collimator to block transmitted rays and
prevent burn-out of the detector.
 Reflected rays are not picked up by the
detector due to the angles involved.
 Diffracted rays at the correct orientation for
the configuration are then collected by the
detector.
School of Science and Technology, Online Counseling Resource…
Applications - 1
Single-crystal X-ray diffraction is most
commonly
used
for
precise
determination of a unit cell, including
cell dimensions and positions of atoms
within the lattice.
 Bond-lengths and angles are directly
related to the atomic positions.
The crystal structure of a mineral is a
characteristic property that is the basis
for
understanding
many
of
the
properties of each mineral.
School of Science and Technology, Online Counseling Resource…
Applications - 2
 New mineral identification, crystal
solution and refinement.
 Determination of unit cell, bondlengths, bond-angles and siteordering.
 Characterization of cation-anion
coordination.
 Variations in crystal lattice with
chemistry.
School of Science and Technology, Online Counseling Resource…
Strengths X-ray Diffraction
 No separate standards required
 Non-destructive
 Detailed crystal structure, including unit cell
dimensions, bond-lengths, bond-angles and
site-ordering information
 Determination of crystal-chemical controls on
mineral chemistry
 With specialized chambers, structures of high
pressure and/or temperature phases can be
determined.
 Powder patterns can also be derived from
single-crystals by use of specialized cameras
(Gandolfi)
School of Science and Technology, Online Counseling Resource…
Limitations X-ray Diffraction
 Must have a single, robust (stable)
sample, generally between 50–250
microns in size.
 Optically clear sample .
 Twinned samples can be handled with
difficulty .
 Data
collection
generally
requires
between 24 and 72 hours
School of Science and Technology, Online Counseling Resource…
Molecular Replacement (MR)-1
 In cases where the crystal under investigation
is isomorphous with the known one (having
the same space group and cell constants
within experimental error), analysis can
proceed
directly
by
difference
Fourier
methods.
 However, more commonly, isomorphism does
not exist, and it becomes necessary to seek
other ways of utilizing the known structure
information to facilitate the target structure
determination.
 To this end, the Molecular Replacement (MR)
method has proved to be particularly
successful
School of Science and Technology, Online Counseling Resource…
Molecular Replacement (MR)-2
The prerequisites for using the MR method are:
 an observed diffraction pattern - intensities - for
the unknown structure, or the target;
 the atomic coordinates of an homologous
protein structure, or the probe.
 The MR task involves positioning the probe
within the unit cell of the target crystal in such a
way that the theoretical diffraction pattern that
would result from this model closely matches
the experimental one.
School of Science and Technology, Online Counseling Resource…
Molecular Replacement (MR)-3
 One
molecule
is
present
in
the
asymmetric unit, six parameters (three
rotational and three translational),
describe how the probe is placed in the
unit cell.
 Study
on
these
six
parameters
determine the position of the probe that
gives the best agreement between
observed
and
calculated
structure
factor.
 This study needs too much computation.
School of Science and Technology, Online Counseling Resource…
Molecular Replacement Method-1
 The first is the determination of the
correct orientation of the probe, and the
second is the determination of the
position
of
the
correctly-oriented
molecule within the unit cell.
 From the theoretical analysis of the
properties of the Patterson function it
became obvious that such a sixparameter search could be reduced to
two three-dimensional problems.
School of Science and Technology, Online Counseling Resource…
Molecular Replacement Method-2
 We have a probe molecule A and
the unknown molecule A' similar to
A.
 The position of A' is different from
A.
 To superimpose the molecule A
with A' we have, firstly, to apply
the rotation R, and then the
translation T.
This figure shows
a
pictorial representation
 Therefore, the main aim of the MR
of the MR problem.
method is to find these two
operators, or, in other terms, to
solve
the
Rotation
and
the
Translation functions.
School of Science and Technology, Online Counseling Resource…
Molecular Replacement Method-3
Factors affecting
 Solving
a
structure
by
the
molecular
replacement method is not always a straight
forward task.
 Depending on the complexity of the problem,
a number of important factors must be taken
into consideration in order to ensure a correct
structure determination.
 The
factors
affecting
the
molecular
replacement solution are crystal and Xray data.
School of Science and Technology, Online Counseling Resource…
Factors affecting MR - 1
The crystal




The crystal form most favorable for the molecular
replacement method is the one with only a single
molecule in the asymmetric unit.
The presence of multiple copies of the protein in the
asymmetric unit reduces the signal-to-noise ratio for the
correct peaks on the rotation and translation function.
Furthermore, the presence of non-crystallographic
symmetry
between
these
copies
can
introduce
difficulties in determining correct solutions.
However, there are many cases in which the structure
was solved by molecular replacement method where
there are several molecules in the asymmetric unit.
School of Science and Technology, Online Counseling Resource…
Factors affecting MR - 2
The X-ray data
 It is very important that the experimental
data are as complete as possible, ideally
100%.
 The data should be of high quality.
 Possible problems include:
 systematically missing regions,
 ice ring,
 detector overloads, etc.
School of Science and Technology, Online Counseling Resource…
Molecular Replacement Search Probe
 The choice of a search probe has to be made
by considering possible structural similarities
with the target molecule (within 1.0 - 1.5 Å
r.m.s.d.).
 The two structures should have, at least,
around 30 - 40% amino-acid sequence identity
and, in general, the higher the better.
 Also, when the choice for the search probe is
given between a crystallographic structure and
an NMR structure, the former, normally,
provides a higher degree of success.
School of Science and Technology, Online Counseling Resource…
Molecular Replacement Search Probe
 This has mainly to do the imprecision of
the NMR protein model.
 Furthermore, the search model does
not necessarily have to be used in its
integrity; some parts of it can be
removed completely
 or the side chains of some residues can
be trimmed to alanine/glycine residues
in the regions where the largest
differences with the unknown structure
are expected.
School of Science and Technology, Online Counseling Resource…
What You Learn…
 You have learnt :
 X-ray scattering techniques.
 Principles
of
Single-crystal
X-ray
Diffraction
 Instrumentation used for Single-crystal Xray Diffraction
 Applications
of
Single-crystal
X-ray
Diffraction Oparins Hypothesis
 Strengths and Limitations of Single-crystal
X-ray Diffraction
School of Science and Technology, Online Counseling Resource…
Critical Thinking Questions
1. Describe a non-destructive analytical
technique
for
describing
crystallographic structure?
2. How to determine complete structure
of crystalline materials?
© 2007, YCMOU. All Rights Reserved.
38
School of Science and Technology, Online Counseling Resource…
Hints For Critical Thinking Question
1. X-ray scattering techniques
2. Single-crystal X-ray Diffraction.
© 2007, YCMOU. All Rights Reserved.
39
School of Science and Technology, Online Counseling Resource…
Study Tips - 1
 Book

Title:Introduction to
Mineral Sciences.

Author: Putnis, A

Cambridge University Press
 Book

Title: ABC Of Biology

Publisher :Holy Faith
 Book

Title: Biological Science

Author: Taylor, Green &
Stout
School of Science and Technology, Online Counseling Resource…
Study Tips - 2
www.en.wikipedia.org
Microsoft Encarta Encyclopedia
•http://www.jiscmail.ac.uk/lists/xrd.html
•Introduction to Xray Diffraction from Links for
Mineralogists, Institute of Mineralogy, University of
Wuerzburg.
•An Introduction to the Scope, Potential and
Applications of X-ray Analysis, from the International
Union of Crystallography
School of Science and Technology, Online Counseling Resource…
End of the Presentation
Thank You !