RAMOT at Tel Aviv University Ltd.

A novel heterologous overexpression system for mammalian membrane protein expression, purification and crystallization

Posted by RAMOT at Tel Aviv University Ltd.Responsive · Innovative Products and Technologies · Israel

Summary of the technology

Cloning and expression methods as well as constructs were developed, that yields significantly higher expression of heterogeneous membrane proteins in bacterial cells, hence facilitating proteins research and crystallization abilities. This is achieved by addition of hydrophilic alpha-helical segments to the desired membrane protein, via a cloning procedure. Two different helices enable construction of eight derivatives of the desired protein. The helices facilitate crystallization by providing the necessary hydrophilic surfaces for crystal formation.
Eight mammalian membrane proteins and two bacterial protein were overexpressed in our bacterial system.
All fusion proteins were solubilized by relatively mild detergents.
All fusion proteins were present in distinct size - monomers or dimers.
All of them were purified standard purification techniques.
The yields of the purified proteins were between 3.6-6.5mg (mammalian) to 3-10mg (bacterial) per liter culture.
All tested proteins were amenable to be concentrated and detergent exchanged for crystallization.
Tested proteins demonstrated both in vitro and in vivo activity
Project ID : 2-2007-22

The Technology

A bacterial cloning and expression system for heterogeneous membrane protein expression (mammalian, bacterial) has been developed. It yields significantly higher expression of heterogeneous membrane proteins in bacterial cells, hence facilitating proteins research and crystallization abilities. This is achieved by addition of hydrophilic alpha-helical segments to the desired membrane protein, via a cloning procedure. Two different helices (hydrophilic bacterial proteins (YaiN (α) and YbeL(β)), enable construction of eight derivatives of the desired protein. The helices facilitate crystallization by providing the necessary hydrophilic surfaces for crystal formation.

Data-to date

10 membrane proteins have been purified (yields: 3.6-6.5mg (mammalian) to 3-10mg (bacterial) per liter culture) including divalent metal ion transporter from E. Coli and mammalian neurotransmitter transporters

The Need

Membranes are the site for the regulation of numerous biological processes and activities; control of these processes is mostly performed by membrane bound proteins. Expression and crystallization of membrane proteins is, however, a formidable task. Over-expression in bacteria encounters problems due to overloading of the membrane and the crystallization process is hampered by the lack of hydrophilic regions crucial for forming inter-molecular bonds in the crystal.

The absence of detailed structural information on membrane proteins severely limits drug discovery efforts and confounds elucidation of numerous mechanisms fundamental to biology. Currently the Pharma industry has to rely on molecular modeling of structures which is often unreliable, hence the great need for reliable structural data that is best derived from crystal structures.

Patent status

Granted patents US, DE, FR, UK

Project manager

Elisha Natan
Director BD

Project researchers

Nathan Nelson
T.A.U Tel Aviv University, Life Sciences
The Department of Biochemistry and Molecular Biology

Shani Leviatan
T.A.U Tel Aviv University, Life Sciences
The Department of Biochemistry and Molecular Biology

Anat Lavi-Itzkovitz
T.A.U Tel Aviv University, Life Sciences
The Department of Biochemistry and Molecular Biology

Related keywords

  • Medicine, Human Health
  • Medical Technology / Biomedical Engineering
  • Biology / Biotechnology
  • Cellular and Molecular Biology Technology
  • Microbiology Technology
  • Genome Research
  • Bioinformatics Technology
  • Micro- and Nanotechnology related to Biological sciences
  • Microbiology Market
  • Micro- and Nanotechnology related to Biological sciences
  • Biochemistry / Biophysics Market
  • Stem cells and biobanks
  • Cellular and Molecular Biology Market
  • Bioinformatics Market
  • Diagnostic
  • Therapeutic
  • Other Medical/Health Related
  • Clinical Medicine
  • Life Sciences and Biotechnology
  • Research / Development tools

About RAMOT at Tel Aviv University Ltd.

Technology Transfer Office from Israel

Ramot is Tel Aviv University's (TAU) technology transfer company and its liaison to industry, bringing promising scientific discoveries made at

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