Barrientos,L.G.; Louis,J.M.; Botos,I.; Mori,T.; Han,Z.Z.; O'Keefe,B.R.; Boyd,M.R.; Gronenborn,A.M.: Structure 10: 673-686, 2002.
Abstract:
Objective: To explore the feasibility of expressing the potent
HIV-inactivating protein, cyanovirin-N (CV-N), in the human commensal bacterium
Streptococcus gordonii as a possible approach for local delivery of CV-N to
prevent sexual transmission of HIV-1.
Design and methods: To express
CV-N in S. gordonii, we used the host-vector system we had previously
developed. CV-N was expressed as a fusion protein both attached to the
bacterial surface and secreted in soluble form in the supernatant of liquid
cultures. The soluble form of recombinant CV-N was tested for gp120-binding
activity in an ELISA experiment, whereas S. gordonii strain expressing CV-N on
the surface was analyzed in an in vitro HIV capturing assay. Results:
Two recombinant S. gordonii strains secreting or displaying CV-N on
the bacterial surface were constructed and the expression of CV-N was confirmed
by immunoblot and cytofluorimetric analysis. The secreted form of recombinant
CV-N exhibited a concentration-dependent binding to envelope glycoprotein gp120
of HIV-1, although with low affinity, whereas CV-N displayed on the bacterial
surface was able to capture HIV virions efficiently.
Conclusion: We
expressed the anti-HIV protein CV-N in S. gordonii in a biologically
active form. This represents a first step in the development of a system to
deliver and maintain an effective concentration of a microbicide in the vaginal
mucosa.
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