High-resolution cryo-electron microscopy study of structure and dynamics of yeast fatty acid synthase by single particle analysis [Elektronische Ressource] / von Preeti Kumari

High-resolution cryo-electron microscopy study of structure and dynamics of yeast fatty acid synthase by single particle analysis. Dissertation zur Erlangung des Doktorgrades der Naturwissenschaften vorgelegt im Fachbereich 14 Biochemie, Chemie und Pharmazie der Johann Wolfgang Goethe Universität in Frankfurt am Main von Preeti Kumari Aus Munger Indien Frankfurt am Main (2009) (D30) 1 CONTENTS ABSTRACT
 8 
CHAPTER
1
 10
INTRODUCTION
 10
1.1
 FATTY
 A CIDS
 10
1.2
 FATTY
 A CIDS
 YNTHESIS
 12
1.3
 TYPE
I
FATTY
 A CIDS
 YNTHASE:
A
MEGASYNTHASE
MACH INE
 18
1.3.
1MAMMALIAN
FATTY
 ACID
SYNTHASE
 18
1.3.
2FUNGAL
FATTY
 ACID
SYNTHASE
 21
1.4
 FAS
INHIBITION
 24
1.5
 A CYLC
 ARRIER
PROTEIN
 26
CHAPTER
2
 28
ELECTRON
MICROSCOP&Y
M
 ETHODS 
 28
2.1
 INTRODUCTION
 28
2.2
 SINGLE
PARTICLE
A NALYSIS
 33 
2.2.

1 IMAGE
FORMATION
 33
2.2.
2NOISE
IN
EM‐IMAGES
 33
2.2.
3CONTRAST
TRANSFER
FUNCTION
 34
2.2.
42D 
IMAGE
ALIGNMENT
AND
CLASSIFICATION
 38
2.2.
53D ‐RECONSTRUCTION
METHODS
 41
2.2.
6RESOLUTION
ASSESSMENT
 46
2.2.
7MAP
 ANALYSIS
 47
2.3

 MATERIALS
AND
METHODS 
 49
2.3.

1 NEGATIVE
STAIN
 49
2.3.

2 CRYO ‐SAMPLE
PREPARATION
 49
2.3.
3ELECTRON
CRYO‐MICROSCOPY
 50
2.3.
4X‐RAY
SOLUTION
SCATTERING
 51
2.3.
5DATA
PROCESSING
 51
2.3.
6RESOLUTION
ASSESSMENT
 52
2.3.
7ESTIMATE
OF
3D ‐VARIANCE
 52
2.3.
8 MAP
VISUALIZATION
AND
ANALYSIS
 53
CHAPTER
3
 54
RESULTS
 54
3.1
 INITIAL
3D ­MAP
 54
3.2
 HIGH
RESOLUTION
3D ­MAP
 60
3.