ELP-FUSION TECHNOLOGY FOR BIOPHARMACEUTICALS - Fusion Protein Technologies for Biopharmaceuticals: Applications and Challenges - page 204

14

ELP-FUSION TECHNOLOGY FOR
BIOPHARMACEUTICALS

D OREEN M. F LOSS , 1 U DO C ONRAD , 2 S TEFAN R OSE -J OHN , 3 AND J ¨ RGEN S CHELLER 1
1 Institute of Biochemistry and Molecular Biology II, Medical Faculty, Heinrich-Heine-University, D¨sseldorf, Germany
2 Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Gatersleben, Germany
3 Institute of Biochemistry, Christian-Albrechts-University, Kiel, Germany

14.1 Introduction
14.2 ELP-based protein purification
14.3 ELPylated proteins in medicine and nanobiotechnology
14.4 Molecular pharming: a new application for ELPylation
14.5 Challenges and future perspectives
14.6 Conclusion
References

component of elastic fibers [3]. The protein is synthesized as
a soluble monomer, tropoelastin, which is subsequently
assembled into a polymeric structure in the ECM and
stabilized by cross-links at Lys residues through the action
of lysyl oxidase [2]. This is actually the property that confers
the conspicuous physical features of the natural elastin
protein [4]. Human, chicken, bovine, and rat tropoelastin
have a significant homology at both DNA and amino acid
levels [5].
Elastic fibers are composed of two entities: amorphous
elastin and a fibrillar component, the microfibrils. During the
formation of the elastic fiber, microfibrils, which are made
up of at least five distinct proteins [6], act as a scaffold, on
which elastin is deposited [4]. Elastin from higher verte-
brates contains over 30% Gly and

14.1 INTRODUCTION

Fusion proteins are successfully used as biopharmaceuticals
and a variety of such therapeutics are investigated in clinical
trials or already reached the market [1]. Beyond the fusion of
effector molecules to Fc domains, albumin or transferrin to
increase the plasma half-life of the resulting therapeutic,
cytotoxicity effects are improved by fusion to toxins,
enzymes, or cytokines. Over the past years, a new fusion
technology based on artificial repetitive polypeptides origi-
nated from mammalian elastin came into the interest of
scientists.
In connective tissues of the large arterial blood vessels,
lung parenchyma, elastic ligaments, and skin, which are
subjected to repetitive and reversible deformation, the major
protein within the extracellular matrix (ECM) is elastin, an
extremely insoluble protein. Furthermore, this structural
protein is also present in some cartilaginous tissues [2].
Elastin is found in all vertebrate species, including cartilag-
inous fish, but not in invertebrates and it is an important

75% of the entire
sequence consists of just four hydrophobic amino acids
(Gly, Val, Ala, and Pro) characteristically present as tetra-
(Val-Pro-Gly-Gly), penta- (Val-Pro-Gly-Val-Gly) or hexa-
repeats (Ala-Pro-Gly-Val-Gly-Val) and forming the hydro-
phobic domain of tropoelastin, which are principally respon-
sible for the elasticity of the protein [7]. In contrast, the
hydrophilic domains of tropoelastin rich in Lys and Ala are
involved in the cross-linking process. These domains often
consist of Lys stretches separated by Ala residues such as
Ala-Ala-Ala-Lys-Ala-Ala-Lys-Ala-Ala [5]. Derivatives of
artificial repetitive polypeptides derived from mammalian
elastin, for example, poly(Val-Pro-Gly-Xaa-Gly), where the
guest residue Xaa is any natural amino acid except proline
[8], have been designated as elastin-like polypeptides
(ELPs) and the process of complementing target proteins
by their C- or N-terminal fusion to an ELP has been termed