title: Flavohemoglobin and nitric oxide detoxification in the human protozoan parasite Giardia intestinalis.
abstract: Flavohemoglobins (flavoHbs), commonly found in bacteria and fungi, afford protection from nitrosative stress by degrading nitric oxide (NO) to nitrate. Giardia intestinalis, a microaerophilic parasite causing one of the most common intestinal human infectious diseases worldwide, is the only pathogenic protozoon as yet identified coding for a flavoHb. By NO amperometry we show that, in the presence of NADH, the recombinant Giardia flavoHb metabolizes NO with high efficacy under aerobic conditions (TN=116+/-10s(-1) at 1microM NO, T=37 degrees C). The activity is [O(2)]-dependent and characterized by an apparent K(M,O2)=22+/-7microM. Immunoblotting analysis shows that the protein is expressed at low levels in the vegetative trophozoites of Giardia; accordingly, these cells aerobically metabolize NO with low efficacy. Interestingly, in response to nitrosative stress (24-h incubation with 5mM nitrite) flavoHb expression is enhanced and the trophozoites thereby become able to metabolize NO efficiently, the activity being sensitive to both cyanide and carbon monoxide. The NO-donors S-nitrosoglutathione (GSNO) and DETA-NONOate mimicked the effect of nitrite on flavoHb expression. We propose that physiologically flavoHb contributes to NO detoxification in G. intestinalis.
authors.affiliation: Department of Biochemical Sciences, CNR Institute of Molecular Biology and Pathology and Istituto Pasteur - Fondazione Cenci Bolognetti, Sapienza University of Rome, I-00185 Rome, Italy.
authors.forename: Daniela
authors.initials: D
authors.lastname: Mastronicola
authors.forename: Fabrizio
authors.initials: F
authors.lastname: Testa
authors.forename: Elena
authors.initials: E
authors.lastname: Forte
authors.forename: Eugenio
authors.initials: E
authors.lastname: Bordi
authors.forename: Leopoldo Paolo
authors.initials: LP
authors.lastname: Pucillo
authors.forename: Paolo
authors.initials: P
authors.lastname: Sarti
authors.forename: Alessandro
authors.initials: A
authors.lastname: Giuffrè
mesh_terms: D000332:Aerobiosis; D016829:Giardia lamblia / Q000187:drug effects / Q000378:metabolism*; D006454:Hemoglobins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D009243:NAD / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D009603:Nitroso Compounds / Q000494:pharmacology; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D026422:S-Nitrosoglutathione / Q000494:pharmacology
chemical_list: D006454:Hemoglobins; D020030:Nitric Oxide Donors; D009603:Nitroso Compounds; D011994:Recombinant Proteins; D009243:NAD; C094210:2,2'-(hydroxynitrosohydrazono)bis-ethanamine; D009569:Nitric Oxide; D026422:S-Nitrosoglutathione