With the aim to better understand the molecular interactions underlying mosquito salivary gland invasion by plasmodium sporozoites, we are displaying on the surface of lambda phages putative adhesive domains from few selected plasmodium proteins which may be involved in the invasion of mosquito salivary glands (domains from TRAP, CS, MAEBL, CRMPs). The lambda system is particularly suitable for our purpose since it makes possible to display a variety of relatively large protein domains (up to 400 aminoacids) as fusions to the C-terminus of the abundant viral capsid protein gpD. Thus, a phage library displaying a collection of plasmodium adhesive domains could be employed for the development of affinity selection experiments targeted to the identification of salivary gland receptors. As a first step toward this direction, adhesive domains from P. falciparum and P. berghei surface proteins putatively involved in recognition/invasion of mosquito salivary glands have been fused to the C-terminus of the lambda phage capsid protein gpD. In order to create the mini-library each domain was amplified by PCR, cloned in a plasmid vector, sequenced and transferred into the lambda vector λD4 (kindly provided by Dr. S. Nasi, University La Sapienza, Rome). After packaging, the resulting libraries were screened for the isolation of recombinant lambda clones. Pilot western blot analysis indicates that, as expected, all the lambda clones identified and isolated to date correctly express the fusion proteins. Screening and verification of correct display of all the selected domains is in progress. The collection of lambda phages will be used for the development of binding assays and used for the affinity selection of salivary gland ligands.

Display of Plasmodium adhesive domains on the surface of lambda phage / Lombardo, Fabrizio; N., Stich; MESTRES SIMON, Montserrat; COLUZZI BARTOCCIONI, Caio Mario; Arca', Bruno. - ELETTRONICO. - (2005). ((Intervento presentato al convegno EMBO Workshop on "Molecular and Population Biology of Mosquitoes and other Disease Vectors" tenutosi a Kolymbari, Crete (Greece) nel 24-31 Luglio 2005.

Display of Plasmodium adhesive domains on the surface of lambda phage.

LOMBARDO, Fabrizio;MESTRES SIMON, montserrat;COLUZZI BARTOCCIONI, Caio Mario;ARCA', Bruno
2005

Abstract

With the aim to better understand the molecular interactions underlying mosquito salivary gland invasion by plasmodium sporozoites, we are displaying on the surface of lambda phages putative adhesive domains from few selected plasmodium proteins which may be involved in the invasion of mosquito salivary glands (domains from TRAP, CS, MAEBL, CRMPs). The lambda system is particularly suitable for our purpose since it makes possible to display a variety of relatively large protein domains (up to 400 aminoacids) as fusions to the C-terminus of the abundant viral capsid protein gpD. Thus, a phage library displaying a collection of plasmodium adhesive domains could be employed for the development of affinity selection experiments targeted to the identification of salivary gland receptors. As a first step toward this direction, adhesive domains from P. falciparum and P. berghei surface proteins putatively involved in recognition/invasion of mosquito salivary glands have been fused to the C-terminus of the lambda phage capsid protein gpD. In order to create the mini-library each domain was amplified by PCR, cloned in a plasmid vector, sequenced and transferred into the lambda vector λD4 (kindly provided by Dr. S. Nasi, University La Sapienza, Rome). After packaging, the resulting libraries were screened for the isolation of recombinant lambda clones. Pilot western blot analysis indicates that, as expected, all the lambda clones identified and isolated to date correctly express the fusion proteins. Screening and verification of correct display of all the selected domains is in progress. The collection of lambda phages will be used for the development of binding assays and used for the affinity selection of salivary gland ligands.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11573/472843
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