Characterization and properties of a 1,3-beta-D-glucan pattern recognition protein of Tenebrio molitor larvae that is specifically degraded by serine protease during prophenoloxidase activation.
Although many different pattern recognition receptors recognizing peptidoglycan and 1,3-beta-D-glucan have been identified in vertebrates and insects, the molecular mechanism of these molecules in the pattern recognition and subsequent signaling is largely unknown. To gain insights into the action mechanism of 1,3-beta-D-glucan pattern recognition protein in the insect prophenoloxidase (proPO) activation system, we purified a 53-kDa 1,3-beta-D-glucan recognition protein (Tm-GRP) to homogeneity from the hemolymph of the mealworm, Tenebrio molitor, by using a 1,3-beta-d-glucan affinity column. The purified protein specifically bound to 1,3-beta-D-glucan but not to peptidoglycan. Subsequent molecular cloning revealed that Tm-GRP contains a region with close sequence similarity to bacterial glucanases. Strikingly, two catalytically important residues in glucanases are replaced with other nonhomologous amino acids in Tm-GRP. The finding suggests that Tm-GRP has evolved from an ancestral gene of glucanases but retained only the ability to recognize 1,3-beta-D-glucan. A Western blot analysis of the protein level of endogenous Tm-GRP showed that the protein was specifically degraded following the activation of proPO with 1,3-beta-D-glucan and calcium ion. The degradation was significantly retarded by the addition of serine protease inhibitors but not by cysteine or acidic protease inhibitor. These results suggest that 1,3-beta-D-glucan pattern recognition protein is specifically degraded by serine protease(s) during proPO activation, and we propose that this degradation is an important regulatory mechanism of the activation of the proPO system.
Zhang
Rong
R
College of Pharmacy, Pusan National University, Jangjeon Dong, Kumjeong Ku, Busan 609-735, Korea.
Cho
Hae Yun
HY
Kim
Hyun Sic
HS
Ma
Young Gerl
YG
Osaki
Tsukasa
T
Kawabata
Shun-ichiro
S
Söderhäll
Kenneth
K
Lee
Bok Luel
BL
D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism*; D004156:Catechol Oxidase / Q000378:metabolism*; D003001:Cloning, Molecular; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000378:metabolism*; D006458:Hemolymph / Q000737:chemistry; D007109:Immunity; D019476:Insect Proteins / Q000378:metabolism; D007814:Larva / Q000737:chemistry; D008969:Molecular Sequence Data; D020539:Sequence Analysis, Protein; D012697:Serine Endopeptidases / Q000378:metabolism*; D015398:Signal Transduction / Q000276:immunology; D013379:Substrate Specificity; D013712:Tenebrio / Q000737:chemistry*
D002352:Carrier Proteins; D004792:Enzyme Precursors; D019476:Insect Proteins; C405332:beta-1,3-glucan recognition protein, insect; C038451:pro-phenoloxidase; D004156:Catechol Oxidase; D012697:Serine Endopeptidases