Title
Evidence for the Regulatory Role of the N-terminal Helix of Secretory Phospholipase A2 from Studies on Native and Chimeric Proteins
Author
Shan Qin, Abhay H. Pande, Kathleen N. Nemec, Xiaomei He, Suren A. Tatulian
Year
2005
Journal
The Journal of Biological Chemistry
Abstract
The phospholipase A2 (PLA2) enzymes are activated by binding to phospholipid membranes. Although the N-terminal α-helix of group I/II PLA2s plays an important role in the productive mode membrane binding of the enzymes, its role in the structural aspects of membrane-induced activation of PLA2s is not well understood. In order to elucidate membrane-induced conformational changes in the N-terminal helix and in the rest of the PLA2, we have created semisynthetic human group IB PLA2 in which the N-terminal decapeptide is joined with the 13C-labeled fragment, as well as a chimeric protein containing the N-terminal decapeptide from human group IIA PLA2joined with a 13C-labeled fragment of group IB PLA2. Infrared spectral resolution of the unlabeled and 13C-labeled segments suggests that the N-terminal helix of membrane-bound IB PLA2 has a more rigid structure than the other helices. On the other hand, the overall structure of the chimeric PLA2 is more rigid than that of the IB PLA2, but the N-terminal helix is more flexible. A combination of homology modeling and polarized infrared spectroscopy provides the structure of membrane-bound chimeric PLA2, which demonstrates remarkable similarity but also distinct differences compared with that of IB PLA2. Correlation is delineated between structural and membrane binding properties of PLA2s and their N-terminal helices. Altogether, the data provide evidence that the N-terminal helix of group I/II PLA2s acts as a regulatory domain that mediates interfacial activation of these enzymes.
Full Article
Instrument
J-810
Keywords
Circular dichroism, Secondary structure, Vesicle interactions, Fluorescence, Ligand binding, Biochemistry