Download Introduction Why biomolecule

RAFT 聚合在生物领域中的应用（一）
Boyer C., Bulmus, V., Davis T. P., Ladmiral V., Liu J., Perrier S. Chemical
Review 2009, 109, 5402-5436.
(作者按姓氏字母顺序排列。本人负责Bioconjugates 部分的编写)
1. Biomolecule-Polymer Conjugates (生物分子和高分子复合物)
Introduction
胚胎干细胞对特定老鼠基因的改造和干扰
Introduction
Why biomolecule-Polymer Conjugates?
Biomolecule-polymer conjugate
Attachment of polymer to biomolecules Improves their stability, solubility and
biocompatibility. It also benefits for the biomolecule’s self-assembly, patterning and
phase behavior.
Maynard, H. D., Heredia K. L. Organic & Biomolecular Chemistry, 2007, 5 (1): 45-53
 Medicine and biotechnology1
1. Vincent, M. J., Duncan, R, Trends in Biotechnology,
2005, 24, 39-47.
 Bio-separations2
 Biosensors3
2. Roy, I., Gupta, M., N., Chemistry and Biology, 2003, 10, 1161-71.
3. Ghica, M. E., Brett, C. M. A., Electroanalysis, 2006, 18, 748-56.
 Diagnosis of diseases4
 Drug delivery5
4. Duncan, R., Nature Reviews of Drug Discovery, 2003, 2, 347-60.
5. Kikuchi, A., Okano, T., Advanced Drug Delivery Review, 2002, 54, 53–77.
 “Graft to”
 “Graft from”
In-situ grafting
Initiator
Introduction
Methods to Prepare Biomolecule-Polymer Conjugates
a) Post-polymerization Conjugation
Polymer
Biomolecule
Biomolecule-polymer conjugate
Drawbacks:
 Multiple steps
 Complicated purification process
 Unavoidable non-specific adsorption
 Multi-site attachment
 Low yield
Introduction
Methods to Prepare Biomolecule -polymer Conjugates
b) In situ polymerization
In situ polymerization
Biomolecule
Biomolecule -polymer conjugate
Advantages
 Less steps
 Less-purification
 Easy to control
 High yield
Introduction
Reversible Addition Fragmentation Chain-Transfer (RAFT)
Polymerisation
a. Atom transfer radical polymerization (ATRP)
 Synthesize well-defined polymeric architectures with predictable MW and
PDI
 Supply a good technique to prepare well-defined protein-polymer
conjugate
b. RAFT polymerization

Polymerize more versatile monomers in more flexible reaction
conditions for a wide range of polymeric architectures

Get rid of the metal catalyst which might complex with some functional
groups on proteins and damage the protein bioactivity.
2. In-situ Synthesis of Protein-polymer Conjugates
-Initiated RAFT
Polymerization
BSA
-S
S
S
S C S
BSA
-S
S
S
S C S
BSA-Polymer Conjugate
J. Liu, V. Bulmus, D. L. Herlambang, C. Barner-Kowollik, M. H. Stenzel,
T. P. Davis, Angew. Chem. Int. Ed. 2007, 46, 3099-3103.
S
O
C
S
O
S
O
C
C
S
S
C
S
S
HS
N
O
H
N
BSA
S
BSA
PEG-A/  irradiation
H2O/DMF
nS
O
O
S
+
S
(2)
BSA
(1)
S
2-pyridinethione
S
O
C
C
S
BSA
S
O
S
(3)
BSA-polymer conjugate
O
8~9
3. Heterotelechelic Protein-Polymer-Biomolecule Conjugates
O
O
N
S
O
S
S
C
S
N
C
C
S
O
S
H2O / DMF
S
BDPET
O
O
N
S
O
S
S
C
BSA
S
PEG-A
C
S
C
S
O
VA-044/250C
S
S
O
O
N
S
O
S
S
C
n
O S
C
S
C
SH
S
O
S
O
Thiocholesterol or thiol terminated
rhodamine B isothiocyanate
SH =
O
n= 8~9
O
O
S
S
O
S
C
n
O S
C
S
C
O
S
S
O
O
n= 8~9
Jingquan Liu, Huiyun Liu, Volga Bulmus, Lei Tao, Cyrille Boyer, Thomas P. Davis,
Journal of Polymer Science Part A: Polymer Chemistry, 2010, 48, 1399-1405.
（IF，3.821） .
2.1 Modification of Bifunctional RAFT Agent
O
C
HO
C
S
S
C
O
O
O
O
Cl
SOCl2
OH
S
C
C
N
C
S
Cl
S
HO
S
S
O
O
N
TEA
CH3CN
S
O
S
S
C
C
N
C
S
S
O
S
S
a
b
c
f
N
S
S
d
O
O
e
C
a
g O
g
S
g
C
C
S
S
O
b
c
N
f
S
e
S
d
g
g
CDCl3
e
f
a
8.5
c,d
b
7.5
6.5
5.5
4.5
ppm
3.5
2.5
1.5
0.5
2.2 Conjugation of Bifunctional RAFT Agent to Protein
O
O
N
S
O
S
S
C
C
N
C
S
S
O
S
S
BSA
SH
O
O
N
H2O / DMF
S
S
O
S
C
C
S
C
S
O
BSA
S
S
1.0
BSA
Modified TRITT
BSA-TRITT conjugation mixture
H
N
Absorption
0.6
6
BSA-TRITT
BSA
S
0.4
RID response
0.8
4
2
0.2
0
0.0
250
300
350
Wavelength/nm
400
450
15
16
17
18
Retention time/min
19
20
2.3 Characterization---- GPC analysis
O
O
N
S
O
S
S
C
C
n
C
S
S
O
BSA
S
O S
O
O
n= 8~9
100
61.7
3.5
61.6
3
60
2
40
1.5
61.4
Response
Ln(M0/M t)
2.5
Conversion
4h polymerization
7h polymerization
10h polymerization
14h polymerization
23h polymerization
61.5
80
61.3
61.2
61.1
1
61.0
20
0.5
0
0
0
5
10
15
Polymerization time/ h
20
25
60.9
60.8
60.7
12
13
14
15
16
17
18
19
20
Retention time/ min
21
22
23
24
25
2.3 Characterization----Cleavage test
O
O
N
S
O
S
S
C
n
C
S
C
SH
O
BSA
O S
HS
O
O
5
n= 8~9
4h polymerization
7h polymerization
10h polymerization
14h polymerization
23h polymerization
3
2.8
100000
MW/ gmol-1
Response
3
120000
2
1
2.6
2.4
80000
2.2
60000
2
1.8
40000
1.6
1.4
20000
1.2
0
0
0
20
40
60
Conversion/ %
26
28
30
32
34
36
38
Retention time/ min
40
42
44
80
1
100
PDI
4
2.4 Characterization----End group check
O
O
N
S
S
S
C
O
n
C
C
S
S
O
BSA
S
O S
O
O
n= 8~9
O
O
Glu
SH
Glu
S
S
O
S
C
n
C
C
S
O
H
N
O
3.5
O
3.0
n= 8~9
BSA-polymer reacted with GSH
BSA-polymer
Absorbance
2.0
1.5
1.0
0.5
0.0
250
300
BSA
S
O S
H2O
2.5
S
350
Wavelength/ nm
400
450
S
3. Summary
 Bifunctional TRITT was successfully modified with pyridyl
disulfide groups on both sides
 The conjugation of bifunctional RAFT revealed that only
one BSA was attached on RAFT
 In-situ polymerization lead to synthesis of bioconjugates
with thiol reactive functional group in one step
 Sometimes research turned out to be much easier?
4. Conjugate of H2NO-KYNPKDKLLY with polymers
as anthrax toxin inhibitor
S
O
S
O
O
Special sequence peptide
O
O
O
Anthrax receptor
O
Polymer chain
O
O
O
Anthrax
N
N
O
peptide
O
P
E
G
This work was done with Prof Heather Maynard at
University of California Los Angeles (UCLA)
Jingquan Liu, Volga Bulmus, Thomas P. Davis, Ronald Li and Heather D Maynard，
Macromolecules, 48, 8-14, 2013. (IF, 5.167)
Tailoring the end groups and cleavage tests
Cholesterol Attachment and cyclodextrIn Complex
*
Eki Setijadi, Lei Tao, Jingquan Liu , Cyrille Boyer, Zhongfan Jia and Thomas P. Davis,
Biomacromolecules, 2009, 10 (9), pp 2699–2707.
Manipulation of the Bioactivity of Glucose Oxidase via RAFT
Controlled Surface Modification
X. Luo, J. Liu*, G. Liu, R. Wang, Z. Liu，J. Polym. Sci. Part A Polym. Chem, 2012，
50, 14, 2786-2793
Micelles/Vesicles as drug carrier