Over-Expression of Membrane Proteins in Bacterial System
The Invention The invention provides for an increase in the expression of heterogeneous membrane proteins as well as crystallization ability. 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. 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. Potential Applications This technology may be applied to a host of membrane proteins and/or protein subunits that have so far proven exceedingly difficult to crystallize. Potential membrane protein candidates include membrane bound receptors, pumps and channels – all of which represent potential therapeutic targets for which detailed structural information is lacking. Advantages The method is general and widely applicable The method is applicable to large proteins (demonstrated for a 66kD protein) The method is inexpensive Induction of protein expression avoids formation of inclusion bodies In contrast to current techniques with antibodies, the hydrophilic helices introduced here are not expected to significantly perturb the resultant structure
Stage The procedure described in the invention was successfully applied to three different proteins: Divalent metal ion transporter from E. Coli, and two neurotransmitter transporters from Mus musculus. A 10-fold increase in expression was found in each case. A neurotransmitter transporter protein has been produced in mg quantities and is planned to enter crystallization studies shortly. Patent Filed Tech Transfer Officer Dr. Nissim Chen Office: +972-3-6406121 Fax: +972-3-6406675 Mail: email@example.com
Type of Offer:
« More Biotech Patents