HEMOGLOBIN BIOSYNTHESIS IN VITREOSCILLA-STERCORARIA DW - CLONING, EXPRESSION, AND CHARACTERIZATION OF A NEW HOMOLOG OF A BACTERIAL GLOBIN GENE

In the strictly aerobic, gram-negative bacterium Vitreoscilla strain C 1, oxygen-limited growth conditions create a more than 50-foId increas e in the expression of a homodimeric heme protein which was recognized as the first bacterial hemoglobin (Hb). The recently determined cryst al structure of Vitreoscilla Hb has indicated that the heme pocket of microbial globins differs from that of eukaryotic Hbs. In an attempt t o understand the diverse functions of Hb-like proteins in prokaryotes, we have cloned and characterized the gene (vgb) encoding an Hb-like p rotein from another strain of Vitreoscilla, V. stercoraria DW. Several silent changes were observed within the coding region of the V. sterc oraria vgb gene. Apart from that, V. stercoraria Hb exhibited interest ing differences between the A and E helices. Compared to its Hb counte rpart from Vitreoscilla strain C1, the purified preparation of V. ster coraria Hb displays a slower autooxidation rate. The differences betwe en Vitreoscilla Hb and V. stercoraria Hb were mapped onto the three-di mensional structure of Vitreoscilla Hb, which indicated that the four changes, namely: Ile7Val, Ile9Thr, Ile10Ser, and Leu62Val, present wit hin the V. stercoraria Hb fall in the region where the A and E helices contact each other, Therefore, alteration in the relative orientation of the A and E helices and the corresponding conformational change in the heme binding pocket of V. stercoraria Hb can be correlated to its slower autooxidation rate. In sharp contrast to the oxygen-regulated biosynthesis of Hb in Vitreoscilla strain C1, production of Hb in V. s tercoraria has been found to be low and independent of oxygen control, which is supported by the absence of a fumarate and nitrate reductase regulator box within the V. stercoraria vgb promoter region, Thus, th e regulation mechanisms of the Hb-encoding gene appear to be quite dif ferent in the two closely related species of Vitreoscilla. The relativ ely slower autooxidation rate of V. stercoraria Hb, lack of oxygen sen sitivity, and constitutive production of Hb suggest that it may have s ome other function(s) in the cellular physiology of V. stercoraria DW, together with facilitated oxygen transport, predicted for earlier rep orted Vitreoscilla Hb.