Sialidases or neuraminidases (EC 188.8.131.52) are enzymes widely distributed in nature, from viruses to vertebrates. They play a key role in the metabolism of sialoglycoconjugates. In particular, they are able to remove sialic acid residues from gangliosides, sialoglycoproteins and sialoligosaccharides. It is becoming more and more evident that the partial desialylation of senescent erythrocyte membrane sialoglycoconjugates is a primary or preliminary signal for erythrophagocytosis. The removal of sialic acid from the sialoglycoconjugates is assumed to be promoted by the sialidases present on the membranes. The presence of two sialidases in human erythrocyte membranes has been previously reported. One acts optimally at acidic pH (4.2-4.7) and the other one at neutral pH. In order to identify the exact nature of these two sialidases and the enzyme–membrane leaflet interaction, we incubated the erythrocyte membranes with different solutions. We obtained interesting results when we treated the membranes with 0.1 M sodium carbonate pH 11.5. In literature, it is reported that this treatment is one of the methods used to discriminate between peripheral and integral cell membrane proteins. We demonstrated that sodium carbonate caused the partial release of the acidic sialidase from erythrocyte membranes, whereas the neutral one remained totally membrane-associated. Our results suggest that the alkaline pH value alone is responsible for the extraction of the acidic enzyme from the membrane and that this protein behaves as a peripheral protein. The analysis of the biochemical and kinetic characteristics allowed us to define the extracted acidic sialidase suitable for purification. Moreover, in order to obtain more information about the nature of the acidic sialidase released from the erythrocyte membranes by sodium carbonate treatment, we performed an analysis of the erythrocyte membranes, on the extracted and non-extractable proteins, by immunoblotting using anti-Neu1 antibody. Our results show the presence of sialidase Neu1 on erythrocyte membranes and the capacity of the sodium carbonate treatment to induce the partial solubilization of Neu1 from the membranes. This represents the first evidence of the presence of Neu1 on human erythrocyte membranes and suggests that the erythrocyte acidic sialidase may be identified with sialidase Neu1.
|Titolo:||Identification and characterization of the acidic sialidase present on human erythrocyte membranes|
|Data di pubblicazione:||2008|
|Parole Chiave:||sialidase ; erythrocyte ; Neu1 ; sialoglycoconjugates|
|Settore Scientifico Disciplinare:||Settore BIO/10 - Biochimica|
|Citazione:||Identification and characterization of the acidic sialidase present on human erythrocyte membranes ; B. Venerando. - Milano : Università degli studi di Milano. DIPARTIMENTO DI CHIMICA, BIOCHIMICA E BIOTECNOLOGIE PER LA MEDICINA, 2008. ((21. ciclo, Anno Accademico 2007/2008.|
|Appare nelle tipologie:||13 - Tesi di dottorato discussa entro ottobre 2010|