FUSION PROTEIN LINKERS: EFFECTS ON PRODUCTION, BIOACTIVITY, AND PHARMACOKINETICS - Fusion Protein Technologies for Biopharmaceuticals: Applications and Challenges - page 55

FUSION PROTEIN LINKERS: EFFECTS ON PRODUCTION,
BIOACTIVITY, AND PHARMACOKINETICS

X IAOYING C HEN , 1 J ENNICA Z ARO , 2 AND W EI -C HIANG S HEN 2
1 Department of Pharmacokinetics, Dynamics, and Metabolism, Pfizer, San Diego, CA, USA
2 Department of Pharmacology and Pharmaceutical Sciences, University of Southern California School of Pharmacy,
Los Angeles, CA, USA

4.1 Introduction
4.2 Overview of general properties of linkers derived from
naturally occurring multidomain proteins
4.3 Empirical linkers in recombinant fusion proteins
4.4 Functionality of linkers in fusion proteins
4.5 Conclusions and future perspective
References

therapeutic effect, and achieve better pharmacokinetic
(PK) and pharmacodynamic (PD) properties than the parent
drugs [7-10]. With the rapid development of protein and
peptide drugs, it is foreseeable that fusion proteins will have
increasing importance in drug development.
Over the years, there has been extensive research about
the construction of various fusion proteins by genetically
fusing functional domains. The rational choice of protein
modules is critical to achieve the desired functions of the
resultant fusion protein. However, another crucial compo-
nent in fusion proteins often overlooked is a suitable linker
moiety to join the protein domains. The direct fusion of
functional domains without a linker may lead to many
undesirable properties, including misfolding of the fusion
proteins [11], low yield in protein expression [12], and
impaired bioactivity [13,14].
This chapter gives an overview of linkers in recombinant
fusion protein technology. First, the properties of linkers in
naturally occurring multidomain proteins are discussed.
The following section introduces the empirical linkers
that have been applied to the successful construction of
recombinant fusion proteins. Readers can refer to these
examples for their specific applications. Finally, the func-
tionalities of linkers in recombinant fusion proteins are
presented to stimulate thoughts about the versatile appli-
cations of linkers. By carefully considering the insertion of
optimal linkers in the design of recombinant fusion proteins,
many improvements, including correct folding [11,12],
enhanced stability [15], and increased biological activities
[14] can be achieved.

4.1 INTRODUCTION

With the advancement of recombinant DNA technology in
the past few decades, genetic fusion of two or more proteins
has created a new class of biological molecules—recombi-
nant fusion proteins. The resultant fusion proteins can obtain
bi- or multifunctional properties derived from each protein
module. There have been numerous applications of fusion
proteins in biomedicine, including development of imaging
techniques by fusing a protein of interest with green fluores-
cent protein (GFP) [1]; improvement of protein expression
and purification by fusing with tags [2], and construction of
bifunctional proteins such as hybrid enzymes [3]. Recom-
binant fusion proteins have also become a new category of
therapeutic agents [4]. They have been widely applied for
drug targeting and delivery, for example, the use of immu-
notoxins in targeted cancer therapy, and antibody-directed
drug delivery across the blood brain barrier (BBB) [5,6].
Additionally, fusing a protein drug with carrier proteins can
significantly prolong the plasma half-life, enhance the