Zocchi Slides

Single molecule sensors for DNA
and proteins
Giovanni Zocchi
Dept. of Physics & Astronomy
UCLA
Nano-technology with soft molecules
Signaling pathway
in the cell
Proteins are nanodevices
Example: conformational change
of the enzyme hexokinase:
Turning the table: molecular deformation
as a probe for binding
Example: a DNA-binding protein
(IHF) bending ds DNA
Micro-mechanical method
to monitor conformation:
- make a single molecular contact
flow
- conformational motion of the contact is
monitored through the displacement of
the bead relative to the slide
- force can be applied on the contact e.g.
through a flow
In practice …
objective
bead
flow
cell
prism
slide
laser beam
DNA hybridization detected from the conformational
change of a single probe oligomer
[Proc. Natl. Acad. Sci. USA 100, 7605 (2003)]
bead
ss
DNA
slide
Hybridization of single 30 base-pairs
oligonucleotide. Bead tethered by single probe.
non-complementary
complementary
pump ON
18
17
length of tether (nm)
pump OFF
16
15
14
13
12
0
5
10
15
time (min)
20
25
Mechanical detection of proteinDNA binding
protein
DNA
IHF protein of E.coli
involved in the integration
of the viral DNA into the
Bacterial DNA
Single IHF molecule
binding and falling off
Two off - rates
→ two modes of binding
Fig.2
32
28
h (nm)
10
24
off
N
20
on
16
1
∆
12
75
76
77
t (min)
78
79
0
50
100
t (s)
150
200
Lifetime of bound state vs. DNA tension
Lifetime (s)
1000
100
10
1
0
2
4
6
Tension (pN)
8
10
12
Sensors for single molecule studies
or
single biomolecules as sensors ?
nano-device performs:
• hybridization assay
• protein-DNA binding assay
New paradigm of molecular detection:
exploits conformational change of the probe
→ single molecule sensitivity with unlabelled targets
Giovanni Zocchi
(“The Boss”)
Jeungphill Hanne
Graduate student
Mukta Singh-Zocchi
Assistant Research Physicist
Awrasa Montrichok
Undergraduate student
Vassili Ivanov
Graduate student
Brian Choi
picture ???
Yan Zeng
Graduate student
Sanhita Dixit
Postdoctoral Fellow