Leishmania (Leishmania) amazonensis:: differential expression of proteinases and cell-surface polypeptides in avirulent and virulent promastigotes

被引:26
作者
Soares, RMD
dos Santos, ALS
Bonaldo, MC
de Andrade, AFB
Alviano, CS
Angluster, J
Goldenberg, S
机构
[1] Univ Brasil, Ctr Ciencias Saude, IMPPG, Dept Microbiol Geral, BR-21941590 Rio De Janeiro, RJ, Brazil
[2] Fundacao Oswaldo Cruz, Dept Bioquim & Biol Mol, Rio De Janeiro, RJ, Brazil
[3] Univ Estado Rio De Janeiro, Fac Ciencias Med, Disciplina Microbiol & Imunol, BR-20551030 Rio De Janeiro, RJ, Brazil
[4] IBMP, BR-81350010 Parana, PR, Brazil
关键词
cell-surface protein; Leishmania (L.) amazonensis; protemases; promastigotes; trypanosomatid; virulence; BSA; bovine serum albumin; CRD; cross-reacting determinant; DTT; dithiothreitol; E-64; trans-epoxysuccinyl L-leucylamido-(4-guanidino) butane; EDTA; ethylenediaminetetraacetic acid; GPI; glycosylphosphatidylinositol; kDa; kilodalton; LPG; lipophosphoglycan; PLC; phospholipase C; PMSF; phenylmethylsulphonyl fluoride; SDS-PAGE; dodecyl sulfate sodium-polyacrylamide gel electrophoresis; VSG; variant surface glycoprotein;
D O I
10.1016/S0014-4894(03)00135-8
中图分类号
R38 [医学寄生虫学]; Q [生物科学];
学科分类号
07 ; 0710 ; 09 ; 100103 ;
摘要
A comparative study of proteolytic enzymes and cell-surface protein composition in virulent and avirulent Leishmania (Leishmania) amazonensis promastigote forms was carried out using one- and two-dimensional dodecyl sulfate sodium polyacrylamide gel electrophoresis (SDS-PAGE). The surface iodinated protein profiles showed two major polypeptides of 65-60 and 50-47 kDa that were expressed in both virulent and avirulent promastigote forms. However, minor quantitative differences were observed in the cell-surface profile between the avirulent and virulent promastigotes. These included polypeptides of 115, 52, 45, 32, and 25 kDa that were preferentially expressed in the virulent forms. Two-dimensional SDS-PAGE showed an accentuated expression of acidic polypeptides; some of them differentially expressed in the promastigote forms analyzed. Live parasites treated with glycosylphosphatidylinositol (GPI)-specific phospholipase C (PLC) from Trypanosoma brucei and immunoprecipitated with the cross-reacting determinant (CRD) antibody recognized three major polypeptides of 65-60, 52, and 50-47 kDa, hence suggesting that these peptides were anchored to the plasma membrane domains through GPI anchor. Moreover, the polypeptides of 65-60 and 52 kDa were also recognized by the gp63 antiserum. Several metalloproteinase activities were similar in both virulent and avirulent promastigote forms, whereas cysteine proteinase activities, sensitive to E-64, were preferentially expressed in virulent promastigotes. These results suggest that cell-surface polypeptides and intracellular cysteine proteinases might play an important role in the virulence of L. (L.) amazonensis. (C) 2003 Published by Elsevier Inc.
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页码:104 / 112
页数:9
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