Structural And Functional Research Of Peptidoglycan Recognition Protein SA(PGRP-SA) Of Different Bees

Innate immune system is the only immunologic barrier in insects.Peptidoglycan recognition protein(PGRPs)is a key pattern recognition receptor in the insect innate immune system.As a short PGRPs,PGRP-SA can bind to bacterial PGN especially for Lys-type PGN and activate the Toll pathway,which triggers the expression and release of antimicrobial peptides to prevent bacterial infection.Drosophila PGRP-SA structure has been solved,and the structure proved the ablilty of binding Lys-PGN and exhibited the important binding amino acids.In current study,we report structures of important bee pollinators Apis mellifera PGRP-SA(Am-PGRP-SA)and Bombus ignitus PGRP-SA(Bi-PGRP-SA)at 1.86 A and 1.41 A resolution.Both Am-PGRP-SA and Bi-PGRP-SA reserved the overall topological structures and disulfide bonds of known PGRP and shared L shape binding groove and some highly conserved groove residues including Ser153(Ser158 in Drosophila)with Drosophila,which are essential for PGN recognition and activation of Toll pathway.However,the area of PGN binding groove and the lined residues exhibited their own characteristics.Moreover,our experiment showed that peptidoglycan recognition protein SAs(PGRP-SAs)from Bombus ignitus(Bi-PGRP-SA)and Apis mellifera(Am-PGRP-SA)as well as Megachile rotundata PGRP-SA(Mr-PGRP-SA)exhibit an intrinsic ability to preferentially bind to the Dap-type PGN from Bacillus subtilis rather than the Lys-type PGN from Micrococcus luteus;this ability is more analogous to the binding exhibited by PGRP-LCx(Dm-PGRP-LCx)and PGRP-SD than to that exhibited by PGRP-SA in Drosophila.Moreover,Bi-PGRP-SA and Am-PGRP-SA share greater sequence identity with Dm-PGRP-LCx than with PGRP-SD and retain several conserved contact residues-His37/His38,His60/His61,Trp66/Trp67,Ala150/Ala151 and Thr151/Thr152.However,the corresponding contact residue Arg85 is not a major anchor residue in bees,e.g.,bumblebees,honeybees and leaf-cutting bees,and residues Thr151/Thr152 and Ser153/Ser154 of Bi-PGRP-SA and Am-PGRP-SA are deduced to be anchor residues based on in silico analysis.In addition,non-conserved residues Asp67 in Bi-PGRP-SA and Mr-PGRP-SA and His68 in Am-PGRP-SA were deduced to be involved in the binding of Dap-type PGNs in bumblebees,honeybees and leaf-cutting bees.We conclude that structure and specificity of PGRP-SAs in bees are more analogous to PGRP-LCx over PGRP-SA of Drosophila.