Structural biological analysis of activation mechanism of hematocyte by ENF peptide family

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Growth-blocking peptide (GBP) is a small insect cytokine with various activities: larval growth regulation, cell proliferation, and stimulation of a class of lepidopteran insect blood cells (plasmatocytes). GBP was initially identified as a 25 amino acid peptide hormone (1-25 GBP), which suppress the growth of host armyworm, found in the hemolymph of the parasitized armyworm. However, in non-parasitized armyworm larvae, 23 amino acid GBP is expressed in insect plasma. Subsequent studies revealed that the parasitization by wasp induced an elongation of the C-terminal amino acid sequence of GBP (1-28 GBP). In this study, we characterized the GBP analogs, which consist of various length of C-terminal region, by comparison of their biological activities and three-dimensional structures. The results indicate that 1-28 GBP exhibits the strongest activity to suppress the larval growth. NMR analysis shows that these peptides have the same tertiary structures basically, ^-which consist of a dis … More ordered N-terminal region, a structured core stabilized by a disulfide bond and an antiparallel β-sheet in aqueous solution. However, the C-terminal region of 1-28 GBP undergoes conformational transition from a random-coiled state to an a-helical state in the presence of dodecylphosphocholine (DPC) micelles. It suggests that the C-terminal region bound to membrane would have an affects on the larval growth activity.We isolated, identified, and determined the solution structure of a novel peptide from lepidopteran, Pseudaletia separata that induces rapid hemocyte aggregation. The isolated peptide from crude integument extract, designated hemocyte-aggregation factor (AGF), was revealed to contain 32 amino acid residues (H_2N-SVQILRCPDGMQMLRSGQCVATTEPPFDPDSY-COOH) by N-terminal amino acid sequencing and 3'-RACE. NMR spectra of ^<15>N-labelled AGF were recorded at 283-313 K and pH 5.0. The full sequential assignment for all non-proline residues was achieved using ^<15>N-edited TOCSY and NOESY spectra. The solution structure was calculated by CYANA and discussed in relevance to the functional feature of AGF. Less