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Biosciences:
Introduction to Mass
Spectrometry and Its Uses in
a Company Like Decode.
Sigurur V. Smrason, Ph.D.
New Technologies Division
Peptides
Steroids
Proteins
Prostaglandins
Oligonucleotides
Acylglycerols
Oligosaccharides
Bile Salts
Fatty Acids
Phospholipids
Phosphoglycerides
Glycophospholipids
Ceramides
Sphingolipids
balances:
0.001g to 1g 0.0001g
Mass
spectrometers:
Basic Concept:
Play Ping-Pong with Molecules
MALDI TOF MS
EI Quad GC/MS
MALDI TOF
(Organic
The
ESI QTOF
(Organic
The
GC/MS
Small
Organic Molecules MS
....and
The
Ionization
Soft
Produces
analyte
Can be either singly charged (MALDI) or
multiply charged (ESI)
Hard
Ionization: EI
Produces
Mass Analyzers
TOF
MS
Greater
Sensitivity
Separation obtained by the ions traveling
at different speeds
Quadrupole
Greater
MS
Selectivity
Seperation obtained by filtering which ion
can reach the detector
Detector
Quadrupole
Quad MS
Selected Examples
Proteins/peptides
Mw= 320.35 Da
min
m/z = 320
m/z
SNP Genotyping
Pinpoint Assay
Non-pinpoint Assay
ddA = 297.2 Da
ddG = 313.2 Da
ddC = 273.3 Da
ddT = 288.2 Da
SNP Genotyping
MALDI TOF MS
Intensity / A.U.
m = 313.0
ddG = 313.2
m/z= 6674.0
m = 297.1
ddA = 297.2
m/z = 6971.1
6600
6800
m/z
m/z = 6987.0
7000
7200
SNP Genotyping
Aquisition
4.7-7
Protein/peptide Analysis
Higher-order structure
elucidation
Protein-protein interactions
Protein-ligand interactions
Modification characterization
Identification
Quantification
Sequencing
Posttranslational Modifications
Intra- versus intermolecular disulfide bridges
protein
cleavage
E
SH
SH
reduction
HS
SH
SH
MALDI MS
MALDI MS
intesity
intesity
peptide mixture
peptide mixture
m/z
m/z
Posttranslational Modifications
Phosporylation identification of peptides
PO3
reflectron MALDI MS
cleavage
m/z
PO3
m = 98
m = 98
PO3
linear MALDI MS
intesity
intesity
PO3
m/z
Posttranslational Modifications
Phosporylation peptide sequencing
PO3
ESI QTOF MS
intesity
PO3
m/z
intesity
m/z
Protein-ligand Interactions
The pH dependence of the Ras-GTP complex
Ras-GTP 19.4 kDa
pH ~ 4.0
intesity
m/z
pH ~ 3.4
m/z
intesity
ESI QTOF MS
intesity
pH ~ 2.8
m/z
X1-X2-X3-X4-X5-X6-...-Xn
PC-X1-X2-X3-X4-X5-X6-...-Xn
X2-X3-X4-X5-X6-...-Xn
PC-X2-X3-X4-X5-X6-...-Xn
X3-X4-X5-X6-...-Xn
PC-X3-X4-X5-X6-...-Xn
X4-X5-X6-...-Xn
PC-X4-X5-X6-...-Xn
...
V G
intesity
m/z
Conclusions
Mass
or
Mass