PURIFICATION AND CLONING OF THE SALIVARY NITROPHORIN FROM THE HEMIPTERAN CIMEX LECTULARIUS

Cimex lectularius and Rhodnius prolixus contain salivary nitric oxide (NO) that may help them to feed on their vertebrate hosts by promoting vasodilation and inhibiting platelet aggregation. Salivary NO is asso ciated with heme proteins (nitrophorins) that store and transport NO f rom the insect salivary glands to the skin of the host. Ln this study, the salivary nitrophorin of Cimex lectularius was purified by DEAE ch romatography and reverse-phase high-performance liquid chromatography, The purified nitrophorin had a molecular mass of 32.9 kDa. The DEAE-p urified hemoprotein was able to bind NO, and this binding shifted the absorption maximum from 388 nm to 438 nm, The ratio of heme to apoprot ein was estimated to be of 1:1. A cDNA clone of 1079 base pairs was se quenced and was found to code for a protein with a molecular mass of 3 1.7 kDa. The clone sequence was in agreement with the internal peptide sequences obtained from the purified protein. Sequencing of the isola ted clone indicates high similarity to several inositol phosphatases; however, no significant similarities emerged when the sequence of C. l ectularius nitrophorin was compared with that of R. prolixus nitrophor in, the only other nitrophorin known in insect saliva. Because C. lect ularius and R. prolixus belong to two different families of Hemiptera that evolved independently to blood feeding, a case is made for the co nvergent evolution of these two insect nitrophorins.