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4.1.3 Thiol-Selecive Chemistries
Thiols derived from Cys side chains (Fig. 4.1) are highly reactive nucleo-
philes. The theoretical p
of Cys lies at 8.8-9.1. At physiological pH, the
Cys is unprotonated and reactive. Based on their highly reactive nature, Cys
side chains in proteins are mostly found engaged in disulfide bonds. Cys
side chains are typically not found in the free reactive form on the solvent-
exposed surface of VNPs. There is only one example, where reactive and
addressable Cys side chains have been reported: Cys side chains have been
identified on the
K
a
interior
solvent-exposed surface of
Cowpea mosaic virus
(CPMV) (Wang
, 2002c, 2003b). Thiols are nevertheless useful groups
to use for bioconjugation; therefore, a range of VNPs have been generated via
genetic modification to add Cys side chains. Cys-added versions of
et al.
Cowpea
chlorotic mottle virus
(CCMV), CPMV,
Flock House virus
(FHV), MS2, Ad, TMV,
and M13, for example, are available (Destito
et al.
, 2009; Khalil
et al.
, 2007b;
Klem
et al.
, 2003; Kreppel
et al.
, 2005; Miller
et al.
, 2007; Peabody, 2003;
Wang
et al.
, 2002b).
Figure 4.3
Reaction scheme for bioconjugation between a thiol and haloacetyl (A)
and maleimide (B), respectively. Figure provided by courtesy of Vu Hong (TSRI, La
Jolla, CA, USA).
A range of chemistries can be applied to thiols. Those that have been
applied to VNPs include reaction with haloacetyl compounds (such as iodo-
or bromoacetamides), coupling with maleimides, binding to gold, and use of
small mercuric reagents. The reaction schemes for coupling haloacetyl- and
maleimide-derivatives to thiols are given in Fig. 4.3; coupling results in the
formation of a stable thioether bond (Aslam & Dent, 1999; Hermanson, 1996).
4.1.4 Tyr-Selecive Chemistries
The aromatic group of Tyr side chains also provides a feasible target for
protein modification. The phenyl group is, however, only modestly reactive,
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