Purification of a peptidoglycan recognition protein from hemolymph of the silkworm, Bombyx mori.
A method was developed for obtaining a homogeneous silkworm hemolymph protein (peptidoglycan recognition protein, PGRP) which has affinity for peptidoglycan and the ability to trigger the prophenoloxidase cascade upon its binding to peptidoglycan. The purified PGRP had a molecular mass of about 19 kDa and is composed of a single polypeptide with an isoelectric point of 6.5. It bound to peptidoglycan in the absence of divalent cation, whereas its binding to beta1,3-glucan and chitin was not detected. N-Acetyl-D-glucosaminyl-(beta1-4)-N-acetylmuramyl-L-alanyl-D-isogluta mine did not inhibit purified PGRP to bind insoluble peptidoglycan, but fragmented soluble peptidoglycan did. PGRP seemed to require peptidoglycan as a possible ligand to keep its glycan portion consisting of at least two or more of the repeating unit. PGRP did not have any detectable lysozyme activity, and its amino acid composition and amino-terminal sequence of 20 amino acid residues were shown to be different from those of silkworm lysozyme. PGRP seems to be a hitherto unknown protein. In the absence of PGRP, the prophenoloxidase cascade in the plasma fraction of hemolymph could not be triggered by peptidoglycan, indicating that some type of activity, capable of activating the cascade, is generated upon their binding. However, the exact nature of this activity is not yet known. The purified PGRP bound to peptidoglycan did not hydrolyze significantly any of the 26 commercially available peptidyl-7-amino-4-methylcoumarins, substrates for various proteases.
Yoshida
H
H
Institute of Low Temperature Science, Hokkaido University, Sapporo, Japan.
Kinoshita
K
K
Ashida
M
M
D000119:Acetylmuramyl-Alanyl-Isoglutamine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D012831:Bombyx / Q000235:genetics / Q000378:metabolism*; D002240:Carbohydrate Sequence; D006458:Hemolymph / Q000378:metabolism*; D008024:Ligands; D008969:Molecular Sequence Data; D010457:Peptidoglycan / Q000378:metabolism*; D011506:Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*
D008024:Ligands; D010457:Peptidoglycan; D011506:Proteins; D000119:Acetylmuramyl-Alanyl-Isoglutamine; C023868:N-acetyl-beta-glucosaminyl-N-acetylmuramyl-alanylisoglutamine