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Production of small, switchable, bispecific antibodies with EF1 and EF2-tagged scFvs

Pérez Canelón, Eduardo (2020) MOBN03 20191
Degree Projects in Molecular Biology
Popular Abstract
Producing small, bispecific antibodies

Proteins are molecules which come in all shapes and sizes, found in every living being. They are translated from the DNA found in the cells of every organism, and every single protein has different characteristics, which allow them to carry out a number of varied functions. Some proteins emit colors, some proteins move ions in the cell, some proteins receive and send signals, and some proteins are even able to target other specific proteins. These last ones are known as antibodies, and the proteins they target are known as antigens; antibodies are key players in the fight against foreign microorganisms such as virus and bacteria. The part of the protein that can bind to an antigen is known as the... (More)
Producing small, bispecific antibodies

Proteins are molecules which come in all shapes and sizes, found in every living being. They are translated from the DNA found in the cells of every organism, and every single protein has different characteristics, which allow them to carry out a number of varied functions. Some proteins emit colors, some proteins move ions in the cell, some proteins receive and send signals, and some proteins are even able to target other specific proteins. These last ones are known as antibodies, and the proteins they target are known as antigens; antibodies are key players in the fight against foreign microorganisms such as virus and bacteria. The part of the protein that can bind to an antigen is known as the variable region, and if an antibody can target two different proteins it is known as a bispecific antibody.

Molecular biologists are able to treat proteins found in nature as building blocks, and combine them and make new useful proteins that can be helpful in the field of medicine, as well as other fields. These are known as protein constructs, and they are made by combining the different DNA sequences which the body translates to proteins, and introducing this DNA sequence into a bacteria. We can then have the bacteria produce whatever protein translated from the DNA we introduce in it, this is known as expression of a protein.

In this project, we are combining the variable regions of two different antibodies, to a molecule known as Calbindin D9k, this molecule consists of two parts, EF1 and EF2, which bind to each other with very high affinity in the presence of calcium. To see if this concept works, we can then make a construct combining EF1 with the variable region of an antibody that targets green fluorescent protein, which as its name suggests is a protein that emits green light, and we can combine EF2 with the HER2 receptor, which is a is present in larger quantities in breast cancer cells. This way, if there is calcium, our bispecific antibody should be able to bind GFP and HER2, and an experiment could be done in which we would see the cells turn green. The purpose of building an antibody this way is that it would also be much smaller than conventional antibodies, and this would facilitate their movement in, for example, cancer tissues.

A lot of work and optimization had to be done in the expression of the constructs, as well as during their purification for their study. In the end, only one of the constructs could be produced, EF1-scFvGFP, but it was determined that both the EF1 and the scFvGFP parts are functional. In order to produce the whole antibody, more studies will be necessary in the future.

Master’s Degree Project in Molecular Biology, 60 cr., 2019-2020
Department of Biology, Lund University

Advisors: Sara Linse and David O’Connell
Department of Chemistry, Lund University and Conway Institute, University College Dublin (Less)
Please use this url to cite or link to this publication:
author
Pérez Canelón, Eduardo
supervisor
organization
course
MOBN03 20191
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
9006983
date added to LUP
2020-03-23 10:06:22
date last changed
2020-03-23 10:06:22
@misc{9006983,
  author       = {{Pérez Canelón, Eduardo}},
  language     = {{eng}},
  note         = {{Student Paper}},
  title        = {{Production of small, switchable, bispecific antibodies with EF1 and EF2-tagged scFvs}},
  year         = {{2020}},
}