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Factors Influencing the Dimer to Monomer Transition of an Antibody Single-Chain Fv Fragment
 

Summary: Factors Influencing the Dimer to Monomer Transition of an Antibody Single-Chain
Fv Fragment
Katja M. Arndt, Kristian M. Mu¨ller, and Andreas Plu¨ckthun*
Biochemisches Institut, UniVersita¨t Zu¨rich, Winterthurerstrasse 190, CH-8057 Zu¨rich, Switzerland
ReceiVed May 6, 1998; ReVised Manuscript ReceiVed July 1, 1998
ABSTRACT: Antibody single-chain Fv (scFv) fragments are able to form dimers under certain conditions,
and the extent of dimerization appears to depend on linker length, antibody sequence, and external factors.
We analyzed the factors influencing dimer-monomer equilibrium as well as the rate of interconversion,
using the scFv McPC603 as a model system. In this molecule, the stability of the VH-VL interaction can
be conveniently varied by adjusting the ionic strength (because of its influence on the hydrophobic effect),
by pH (presumably because of the presence of titratable groups in the interface), and by the presence or
absence of the antigen phosphorylcholine, which can be rapidly removed due to its very fast off-rate. It
was found that the monomer is the thermodynamically stable form with linkers of 15 and 25 amino acids
length under all conditions tested (35 µM or less). The dimer is initially formed in periplasmic expression,
presumably by domain swapping, and can be trapped by all factors which stabilize the VH-VL interface,
such as the presence of the antigen, high ionic strength, and pH below 7.5. Under all other conditions,
it converts to the monomer. Predominantly monomer is obtained during in vitro folding. Monomer is
stabilized against dimerization at very high concentrations by the same factors which stabilize the VH-
VL interaction. These results should be helpful in producing molecules with defined oligomerization
states.

  

Source: Arndt, Katja - Institut für Biologie III, Albert-Ludwigs-Universität Freiburg
Müller, Kristian - Institut für Biologie III, Albert-Ludwigs-Universität Freiburg

 

Collections: Biology and Medicine; Biotechnology