Introduction

Our research program is devoted to the study of gaseous macro-ions appliedwithin the context of mass spectrometry to challenging analysis problems.

Our work involves instrument development, ion chemistry studies, and applications to analytical problems. This poster summarizes our recent activities in these areas.

Instrumentation

Finnigan (ITMSTM) dual source ion trap mass spectrometer

Finnigan (ITMSTM) 4-source ion trap mass spectrometer

Modified Q STAR quadrupole time-of-flight mass spectrometer

Two modified Q TRAP linear ion trap mass spectrometers

Multi-Source linear ion trap

Cold-ion spectroscopy tandem mass spectrometer

The 4-Source Ion Trap

Glow discharge

source

Ion trap

Transfer optics

(tube lens / linear quad)

Turning quad

ESI / Glow

Discharge

Source

Turbo pump

400 L/s

Turbo pump

400 L/s

-ESI

source

+ESI

source

Modified Dueling Source 3D Ion Trap

ASGDI Source

ESI Source

DC Pulse

Generator

Custom Switch Box

RF Power

Supply

Frequency

Synthesizer

Amplifier

TTL1

Out

Finnigan Electronics

+V -V

500 mTorr

1 mTorr

Quadrupole Ion TrapCEM

Turning

Quad

ESI Tip

Electrostatic

Lenses

Glow

Discharge

Multi-Source Linear Ion Trap

Cold-Ion Spectroscopy Tandem Mass

Spectrometer

Joint project with the

Laboratory of

Timothy Zwier

Source A

(ESI)

Source B

(Glow Discharge)

4K Cold Trap

Detector

Laser Pulse

Leak-in Setup LIT ion/ion Reactor TOF Analyzer

Modified Q Star Quadrupole

Time-of-Flight

Curtain

Plate

Orifice

PlateCurtain

Plate

Orifice

Plate

Q0 Q1 Q2 Q3

Aux AC

Orifice

Skimmer IQ1 ST IQ2 IQ3Exit lens

Deflector

Detector

N2 CAD Gas

Modified QTRAP Linear Ion Trap

RF

Pulsed Triple

Ionization Source

Analyte inlet

Reagent inlet

Reagent inlet

Modified MALDI-QTRAP4000

Linear Ion Trap

Laser Beam

Custom Components

High power rf supplies• Able to drive LITs, QIT,

and mass resolving Quadrupoles

Low power rf supplies• Durable tube-based

design for ion optical devices

• Scanable transistor supplies for simple mass analysis

Ion funnel• Small, simple design

• Significant signal gains on homebuilt interface

• Robust interface

Chemistry

Unimolecular Dissociation Reactions• Peptides• Proteins• Nucleic Acids• Carbohydrates

Ion/Ion Reactions• Proton Transfer (PT)• Electron Transfer (ET)• Specific Covalent Chemistry

Ion/Molecule Reaction• Treatment of Electrosprayed Droplets

e-e-e-

Reagent anion

H2N—CH—C—NH—CH—C—NH—CH—COH

R1 O R2 O R3 O

— — ——— —— ——

y2

b1

z2

c1

x2

a1

y1

b2

z1

c2

x1

a2

H2N—CH—C—NH—CH—C—NH—CH—COH

R1 O R2 O R3 O

— — ——— —— —— —— —— ——

y2

b1

z2

c1

x2

a1

y1

b2

z1

c2

x1

a2

Excitation increases ion displacement from the trap center, whereby

ions undergo increases in kinetic energy. Collisions with the bath gas

convert kinetic energy to internal energy (slow heating).

Less sequence dependence than CID (wider sequence coverage)

Does not cleave labile post translational modifications

H2N—CH—C—NH—CH—C—NH—CH—COH

R1 O R2 O R3 O— — ——— —— ——

y2

b1

z2

c1

x2

a1

y1

b2

z1

c2

x1

a2

H2N—CH—C—NH—CH—C—NH—CH—COH

R1 O R2 O R3 O— — ——— —— —— —— —— ——

y2

b1

z2

c1

x2

a1

y1

b2

z1

c2

x1

a2

Electron Transfer Dissociation (ETD)

Collision-Induced Dissociation (CID)

Electron Transfer Reaction between Azobenzene

and Disulfide-linked Peptide [M+3H]3+

700 702698

200 400 600 800 1400m/z

0

100

Rel

ativ

e A

bu

nd

ance

, %

Az1+●

[B]+●

ABz2+●

1000 1200

Ac1+

Bc1+

Bc2+

3+

Bc3+

Bc4+

2+[B-S-H]+

1+NH2

O

(CH2)3N

CH3

H3C

H3C

+_[A-SH2]+

[A+S]+●

[B+S+H]+

[A]+

[A-2H]+

646 648644[B+H]+

Pep V: (q)A G C K(q)-OMe

(q+)T F T S C-OMe

S

S

A: A-Chain

B: B-Chain

DC CID: Fragmentation vs Time

DC CID: A Broadband Application

Top: DC CID of bromocriptine. 40 VDDC, 7.5 ms,

low mass cutoff = 117 m/z.

Middle: DC CID of YGGFL. 12.8 VDDC, 10 ms,

low mass cutoff = 51 Th.

Bottom: DC CID of ubiquitin 8+. 76 VDDC, 10 ms,

low mass cutoff = 234 Th.

100

*(A6–3H)3–

[CuII(A6–3H)]–

(A6–3H)2–•

(A6+O2–3H)2–•

w1–R

ela

tive

Ab

un

da

nce

500 1300300 1100 1500700 1700900 1900100

a) (A6–3H)3– + [CuII(phen)2]2+

b) (A6–3H)3– + [CoII(phen)2]2+

100

500 1300300 1100 1500700 1700900 1900100

(A6–3H)3–

(x 6.2)

(A6–3H)2–•

(A6+O2–3H)2–•

Re

lative

Ab

un

da

nce

#

#

w1–

[CoII(A6–3H)]–*~

c) (A6–3H)3– + [FeII(phen)2]2+

100

500 1300300 1100 1500700 1700900 1900100

(A6–3H)2–•

(A6–3H)3–

(x 6.1)

[FeII(A6–3H)]–

(A6+O2–3H)2–•

Re

lative

Ab

un

da

nce

#

*~

m/z

100

*(A6–3H)3–

[CuII(A6–3H)]–

(A6–3H)2–•

(A6+O2–3H)2–•

w1–R

ela

tive

Ab

un

da

nce

500 1300300 1100 1500700 1700900 1900100

a) (A6–3H)3– + [CuII(phen)2]2+

b) (A6–3H)3– + [CoII(phen)2]2+

100

500 1300300 1100 1500700 1700900 1900100

(A6–3H)3–

(x 6.2)

(A6–3H)2–•

(A6+O2–3H)2–•

Re

lative

Ab

un

da

nce

#

#

w1–

[CoII(A6–3H)]–*~

c) (A6–3H)3– + [FeII(phen)2]2+

100

500 1300300 1100 1500700 1700900 1900100

(A6–3H)2–•

(A6–3H)3–

(x 6.1)

[FeII(A6–3H)]–

(A6+O2–3H)2–•

Re

lative

Ab

un

da

nce

#

*~

100

*(A6–3H)3–

[CuII(A6–3H)]–

(A6–3H)2–•

(A6+O2–3H)2–•

w1–R

ela

tive

Ab

un

da

nce

500 1300300 1100 1500700 1700900 1900100

a) (A6–3H)3– + [CuII(phen)2]2+

100

*(A6–3H)3–

[CuII(A6–3H)]–

(A6–3H)2–•

(A6+O2–3H)2–•

w1–R

ela

tive

Ab

un

da

nce

500 1300300 1100 1500700 1700900 1900100

100

*(A6–3H)3–

[CuII(A6–3H)]–

(A6–3H)2–•

(A6+O2–3H)2–•

w1–R

ela

tive

Ab

un

da

nce

500 1300300 1100 1500700 1700900 1900100 500 1300300 1100 1500700 1700900 1900100

a) (A6–3H)3– + [CuII(phen)2]2+

b) (A6–3H)3– + [CoII(phen)2]2+

100

500 1300300 1100 1500700 1700900 1900100

(A6–3H)3–

(x 6.2)

(A6–3H)2–•

(A6+O2–3H)2–•

Re

lative

Ab

un

da

nce

#

#

w1–

[CoII(A6–3H)]–*~

b) (A6–3H)3– + [CoII(phen)2]2+

100

500 1300300 1100 1500700 1700900 1900100

(A6–3H)3–

(x 6.2)

(A6–3H)2–•

(A6+O2–3H)2–•

Re

lative

Ab

un

da

nce

#

#

w1–

[CoII(A6–3H)]–*~100

500 1300300 1100 1500700 1700900 1900100 500 1300300 1100 1500700 1700900 1900100

(A6–3H)3–

(x 6.2)

(A6–3H)2–•

(A6+O2–3H)2–•

Re

lative

Ab

un

da

nce

#

#

w1–

[CoII(A6–3H)]–*~

c) (A6–3H)3– + [FeII(phen)2]2+

100

500 1300300 1100 1500700 1700900 1900100

(A6–3H)2–•

(A6–3H)3–

(x 6.1)

[FeII(A6–3H)]–

(A6+O2–3H)2–•

Re

lative

Ab

un

da

nce

#

*~c) (A6–3H)3– + [FeII(phen)2]

2+

100

500 1300300 1100 1500700 1700900 1900100

(A6–3H)2–•

(A6–3H)3–

(x 6.1)

[FeII(A6–3H)]–

(A6+O2–3H)2–•

Re

lative

Ab

un

da

nce

#

*~100

500 1300300 1100 1500700 1700900 1900100 500 1300300 1100 1500700 1700900 1900100

(A6–3H)2–•

(A6–3H)3–

(x 6.1)

[FeII(A6–3H)]–

(A6+O2–3H)2–•(A6+O2–3H)2–•

Re

lative

Ab

un

da

nce

#

*~

m/z

Ion/Ion Reactions of Transition Metal

Complex Cations with Multiply Charged

Oligodeoxynucleotide Anions

a) (A6–3H)3– + [CuII(phen)3]2+

500 1300300 1100 1500700 1700900 1900100

# [CuII(A6–3H)]–

(A6–3H)3–

(x 9.1)

(A6–3H)2–●

Re

lative A

bundance

*100 ~

500 1300300 1100 1500700 1700900 1900100

100

b) (A6–3H)3– + [CoII(phen)3]2+

[CoII(A6–3H)]–

(A6–3H)3–

(x 1.8)

(A6–3H)2–●

Re

lative A

bundance

*~

100

500 1300300 1100 1500700 1700900 1900100

(A6–3H)3–

(x 4.3)

c) (A6–3H)3– + [FeII(phen)3]2+

(A6–3H)2–●

[FeII(A6–3H)]–

#

Re

lative A

bundance

*~

m/z

a) (A6–3H)3– + [CuII(phen)3]2+

500 1300300 1100 1500700 1700900 1900100

# [CuII(A6–3H)]–

(A6–3H)3–

(x 9.1)

(A6–3H)2–●

Re

lative A

bundance

*100 ~

a) (A6–3H)3– + [CuII(phen)3]2+

500 1300300 1100 1500700 1700900 1900100

# [CuII(A6–3H)]–

(A6–3H)3–

(x 9.1)

(A6–3H)2–●

Re

lative A

bundance

*100 ~

500 1300300 1100 1500700 1700900 1900100

# [CuII(A6–3H)]–

(A6–3H)3–

(x 9.1)

(A6–3H)2–●

Re

lative A

bundance

*100 ~

500 1300300 1100 1500700 1700900 1900100

100

b) (A6–3H)3– + [CoII(phen)3]2+

[CoII(A6–3H)]–

(A6–3H)3–

(x 1.8)

(A6–3H)2–●

Re

lative A

bundance

*~

500 1300300 1100 1500700 1700900 1900100

100

b) (A6–3H)3– + [CoII(phen)3]2+

[CoII(A6–3H)]–

(A6–3H)3–

(x 1.8)

(A6–3H)2–●

Re

lative A

bundance

*~

100

500 1300300 1100 1500700 1700900 1900100

(A6–3H)3–

(x 4.3)

c) (A6–3H)3– + [FeII(phen)3]2+

(A6–3H)2–●

[FeII(A6–3H)]–

#

Re

lative A

bundance

*~

m/z

100

500 1300300 1100 1500700 1700900 1900100

(A6–3H)3–

(x 4.3)

c) (A6–3H)3– + [FeII(phen)3]2+

(A6–3H)2–●

[FeII(A6–3H)]–

#

Re

lative A

bundance

*~

m/z

m/z

m/z

Inte

nis

ty

(Arb

. U

nits)

m/z

Inte

nsity

(Arb

. U

nits)

pdA

40

pdA

41

pdA

42

pdA

43

pdA

44 pdA

45

pdA

46

pdA

47

pdA

48

pdA

49

pdA

50

pdA

51

pdA

52

pdA

53

pdA

54

pdA

55

pdA

56

pdA

57

pdA

58

pdA

59

pdA

60

ESI of a DNA Mixture, pd(A)40-60

Proton Transfer Charge Reduction of Multiply-

Charged pd(A)40-60 Anions

Ion Parking

Mass Spectra of Porcine Elastase Acquired in

(a) Pre Ion/Ion, (b) Post Ion/Ion and (c) Ion Parking Modes

30000

20000

15000

10000

5000

25000

1000 2200 300018001400 2600

m/z

Abunda

nce

[M+19H]19+

[M+15H]15+

[M+11H]11+

[M+14H]14+

[M+17H]17+

[M+21H]21+ [M+12H]12+

[M+10H]10+

a)

b)

c)

Charge Inversion Process

2+

+ 2H+

+

+ H+

-

- H+

N

NH

O

OHO

HO

N

HN

O

O

HO

HO

N

NH

O

OOH

OH

N

HN

O

O

OH

OHOO

OO

NN

n-

Peptide Ion

(Bradykinin)

-

- H+

Polyamidoamine (PAMAM)

Dendrimer Anion

1,4-Diaminobutane (DAB)

Dendrimer Cation

m+

N

N

NH2N

H2N

NH2N

H2N

NH2N

H2N

N

H2N

H2N

N

N

N

N

NH2N

H2N

NH2N

H2N

NH2N

H2NH2N

H2N

N

N

N

NNH2

NH2

NNH2

NH2

NNH2

NH2

N

NH2

NH2

N

N

N

NNH2

NH2

NNH2

NH2

NNH2

NH2

N

NH2

NH2

N

NN

Sequential Information via Covalent Modification

made in the Gas Phase

[M+♦]-

b♦6

b♦5

b♦4

b♦3

b♦1

y♦7

a)9e6

6e6

3e6

1.6e8

8e7

200 400 600 800 1000 12000

0

[M-H]-

y7

b)

D R V Y I H P F

[FBDSA-H]-

[M+♦-H2O]-A

bu

nd

ance

(A

rb.

Units)

m/z

CID of unmodified angiotensin

CID of modified angiotensin with

4-formyl-benzene disulfonic acid

Inter-Molecular Cross-Linking in the Gas-Phase

m/z

300 500 900 1100 1300 1700100 700 1500

0

6.2x E6

300 500 900 1100 1300 1700100 700 1500

0

1.4x E6

Re

lative

Ab

unda

nce

(A

rb. U

nit)

x2.5

3.1x E6

7.0x E5

(a)

(b)

-(sulfo-NHS)

[M1+BS3-2Na-(sulfo-NHS)+M2+2H]2+

[M1+M2+BS3-2Na-2(sulfo-NHS)+2H]2+

-H2O

-H2O

b9*2+

b6*

y6*

b5*

y5*b4*

y4*

b3*

y3*

b9

b2*

y2*

b2b3 b4 b5 b6

b9*2+

y9*2+

b1*

y1*

b8*2+

y8*2+b7*

2+

y7*2+

Ion Trap CID of [YGGFLK+BS3-2Na-(sulfo-NHS)+KKKKKKKKKK+2H]2+

MS3 of [YGGFLK+BS3-2Na-2(sulfo-NHS)+KKKKKKKKKK+2H]2+

* denotes KKKKKKKKKK fragment ions cross-linked to YGGFLK

Charge Inversion/ETD of a Phosphopeptide

-

- H+

7+N

N

NH2N

H2N

NH2N

H2N

NH2N

H2N

N

H2N

H2N

N

N

N

N

NH2N

H2N

NH2N

H2N

NH2N

H2NH2N

H2N

N

N

N

NNH2

NH2

NNH2

NH2

NNH2

NH2

N

NH2

NH2

N

N

N

NNH2

NH2

NNH2

NH2

NNH2

NH2

N

NH2

NH2

N

NN

-1 Phosphopeptide Anion

2+

+ 2H+

+2 Phosphopeptide Cation

+7 DAB Dendrimer G4 Cation

+Charge Inversion

+

-1 Azobenzene Anion

-

ETD

Ab

un

da

nce

(A

rb.

Un

its)

+Na

+2Na+3Na

[M+3H]3+

[M+3H]3+

+Na

+3Na+4Na+5Na

+6Na+7Na

+2Na+8Na

+9Na+10Na

[M+3H]3+

+Na+2Na+3Na

+4Na+5Na

+6Na

Inte

nsity (

Arb

. U

nit)

(a)

(b)

(c)

+nESI

No Leak

HCl Leak

TFA Leak

(e)

(d)

(f)

+KNa +3Na

-nESI

No Leak

Acetic Acid

Leak

Formic Acid

Leak

[M-4H]4-

+K

+K

+Na

+KNa +3Na

[M-4H]4-

+Na

+Na

[M-4H]4-

Acid Vapor Introduction for Removal of Metal Counter-

Ions of Various DNA 12mers

m/z

+6 -5

+9-7

+16 -10

Cytochrome c100% H2O

BaseLeak-in

BaseLeak-in

AcidLeak-in

AcidLeak-in

Protein Folding

Protein Unfolding

m/z

Inte

nsity (

Arb

. U

nit)

Applications

Protein Mixture Analysis

Database-Assisted Protein Identification

Post-Translational Modification (PTM)

Analysis

―Top-Down‖ Protein Identification

+

+

+ ++

+

+

+

++

+

+++

++

+

+++++

+

++

+

+

+

++

Protein Ionization & accumulationPrecursor ion purification and

concentration via ―ion parking‖

ESI Ion/Ion

chemistry

Ion/Ion

chemistry

Charge state

reduction

m/z

Abundan

ce

4000 5000 6000 7000 8000300020001000

1500

m/z

Abundan

ce

4000 5000 6000 7000 8000300020001000

1500

Dissociation

++

+ +++

+

m/z

Abundan

ce

4000 5000 6000 7000 8000300020001000

1500

m/z

Abundan

ce

4000 5000 6000 7000 8000300020001000

1500

Protein

Identification

Protein Database

search

Relative Informing Power of ESI Based

Top-down Approaches

m/z

+10+20

+30

m/z

Post ion/ion

+1+1

+1

ESI

High resolution

m/z

Modified DNA Sequencing

0

500

1000

1500

2000

2500

250 450 650 850 1050 1250

0

50

100

150

200

250

300

350

400

450

500

250 450 650 850 1050 1250

ITCID Ps 2’-OMe RNA 6-mer -3 120450Hz_180mV

w1-

x1-

y1-

c1-

b32-

x32-

c32-

d53- x5

3-

[M-AH]3-

y42-

c2-

z42-c4

2-x2

- a5--B b5

3-b3

-[M-AH]2-

y3-

x3-

b1+

a1+

y1+

#

c1+

b32+

a2+-B

y32+

c32+

#

#

#

#

z2+

y2+

c2+

y42+

w2+

#y5

2+ x52+

[M-GH]2+

[M-AH]2+

[M-CH]2+

y3+ x3

+

ITCID Ps 2’-OMe RNA 6-mer +3 120088Hz_250mV

# - internal fragment

O

HO

O OMe

A

P

O

O SH

O

O OMe

C

P

O

O OH

O

O OMe

C

P

O

O SH

O

O OMe

G

P

O

O OH

O

O OMe

A

P

O

O SH

O

OH OMe

G

Abundance (

Arb

. U

nits)

m/z

Post Ion-Ion Reaction MS/MS Spectra of the

[M + 9H]9+ Ion of α-synucleinA

bun

da

nce

9000 10000 11000 12000 13000 14000 150000

20

100

120

60

80

40

[M+H]+

b140+

b139+

b134+-b137

+/

b134*-b137*

y134+

b130+-b133

+/

b131*/y131+

b121+ - b127

+/

b121*,b122*,/

b125*-b127*/

y119+- y120

+/

y122+ - y124

+

y117+

b119+/

b119*

b116+ - b118

+/

b115*, b116*,b118*/

y114+ - y115

+

b109+ - b114

+/

b109*,b110*,

b112* - b114*/

y106+ - y108

+, y110+b107

+/

b107*

y102+

b98+/

b98*b92

+

y91+

b95+

y90+

b97+

y94+

b105+/

b105*

y95+

b104+/

b104*

b103+

b102+

y97+

b100+

500

1000 1500 2000 2500 3000 3500

204060

140120

200180

0

160

10080

y3+

b3+y4

+

b4+

y5+

b5+

b6+ - b10

+/

y7+ - y9

+ y10+

b11+

y11+

b12+

b13+

y13+

y14+

b15+

b16+

y15+

b17+

b18+

b19+/

y16+

y17+

b20+

b21+

y19+

b22+

b23+

y21+

b24+

y22+

b26+

y24+

b28+/

y25+

b34+

b33+

b32+

b31+

b30+

b29+

5000 6000 70004000 90008000

100

20

120

60

40

0

80

b38+ b52

+

b1192+

+2 speciesb60

+-b62+/

b60*-b62*/

y60+

b34+

b35+

b37+

y33+

b39+

b40+

b41+

b42+,

b42*

b43+

b44+

b45+

y42+

b46+,

b46*

b47+

b48+

b49+

b50+ b53

+

y49+,

y49*

b55+,

b56*

b56+

[M+2H]2+

b66+/

b66*

b63+

y61+

b64+/

b64*

b65+/

b65* b67+ - b71

+/

y64+,y66

+,

y68+,y68*

y69+,

b72+

b73+ - b76

+/

b73*, y70+,

y74+, y74*

b77+ - b83

+/

b81*- b83*/

y79+,y80

+b84

+ - b91+,

b87*/y81+

m/z

From N-terminus to C-terminus:

Scott A. McLuckey• Ken Chanthamontri• Corrine DeMuth• Christine Fisher• Yang Gao• Josh Gilbert• Kerry Hassell• Ryan Hilger• Anastasia Kharlamova• Carl Luongo• Marija Mentinova• William McGee• Boone Prentice• James Redwine• Jessica Espy• John Stutzman• Ian Webb

“Macro-Ionophiles”

COOH

NH3+