Alpha-glucosidase promotes hemozoin formation in a blood-sucking bug: an evolutionary history.
BACKGROUND
Hematophagous insects digest large amounts of host hemoglobin and release heme inside their guts. In Rhodnius prolixus, hemoglobin-derived heme is detoxified by biomineralization, forming hemozoin (Hz). Recently, the involvement of the R. prolixus perimicrovillar membranes in Hz formation was demonstrated.


RESULTS
Hz formation activity of an alpha-glucosidase was investigated. Hz formation was inhibited by specific alpha-glucosidase inhibitors. Moreover, Hz formation was sensitive to inhibition by Diethypyrocarbonate, suggesting a critical role of histidine residues in enzyme activity. Additionally, a polyclonal antibody raised against a phytophagous insect alpha-glucosidase was able to inhibit Hz formation. The alpha-glucosidase inhibitors have had no effects when used 10 h after the start of reaction, suggesting that alpha-glucosidase should act in the nucleation step of Hz formation. Hz formation was seen to be dependent on the substrate-binding site of enzyme, in a way that maltose, an enzyme substrate, blocks such activity. dsRNA, constructed using the sequence of alpha-glucosidase gene, was injected into R. prolixus females' hemocoel. Gene silencing was accomplished by reduction of both alpha-glucosidase and Hz formation activities. Insects were fed on plasma or hemin-enriched plasma and gene expression and activity of alpha-glucosidase were higher in the plasma plus hemin-fed insects. The deduced amino acid sequence of alpha-glucosidase shows a high similarity to the insect alpha-glucosidases, with critical histidine and aspartic residues conserved among the enzymes.


CONCLUSIONS
Herein the Hz formation is shown to be associated to an alpha-glucosidase, the biochemical marker from Hemipteran perimicrovillar membranes. Usually, these enzymes catalyze the hydrolysis of glycosidic bond. The results strongly suggest that alpha-glucosidase is responsible for Hz nucleation in the R. prolixus midgut, indicating that the plasticity of this enzyme may play an important role in conferring fitness to hemipteran hematophagy, for instance.
Mury
Flávia Borges
FB
Laboratório de Química e Função de Proteínas e Peptídeos, Universidade Estadual do Norte Fluminense, Campos dos Goytacazes, Rio de Janeiro, Brazil.
da Silva
José Roberto
JR
Ferreira
Ligia Souza
LS
dos Santos Ferreira
Beatriz
B
de Souza-Filho
Gonçalo Apolinário
GA
de Souza-Neto
Jayme Augusto
JA
Ribolla
Paulo Eduardo Martins
PE
Silva
Carlos Peres
CP
do Nascimento
Viviane Veiga
VV
Machado
Olga Lima Tavares
OL
Berbert-Molina
Marília Amorim
MA
Dansa-Petretski
Marilvia
M
D000818:Animals; D001665:Binding Sites; D002384:Catalysis; D019143:Evolution, Molecular; D005260:Female; D005786:Gene Expression Regulation; D006418:Heme / Q000737:chemistry; D006420:Hemeproteins / Q000737:chemistry*; D006454:Hemoglobins / Q000737:chemistry; D006868:Hydrolysis; D007313:Insecta; D007413:Intestinal Mucosa / Q000378:metabolism*; D008871:Microvilli / Q000378:metabolism; D012330:RNA, Double-Stranded / Q000737:chemistry; D012239:Rhodnius / Q000378:metabolism; D000520:alpha-Glucosidases / Q000737:chemistry*
D006420:Hemeproteins; D006454:Hemoglobins; D012330:RNA, Double-Stranded; C020753:hemozoin; D006418:Heme; D000520:alpha-Glucosidases