Infestin, a thrombin inhibitor presents in Triatoma infestans midgut, a Chagas' disease vector: gene cloning, expression and characterization of the inhibitor

Infestin, a thrombin inhibitor presents in Triatoma infestans midgut, a Chagas' disease vector: gene cloning, expression and characterization of the inhibitor

Author Campos, ITN Google Scholar
Amino, R. Google Scholar
Sampaio, CAM Google Scholar
Auerswald, E. A. Google Scholar
Friedrich, T. Google Scholar
Lemaire, H. G. Google Scholar
Schenkman, S. Google Scholar
Tanaka, A. S. Google Scholar
Institution Universidade Federal de São Paulo (UNIFESP)
Klinikum Univ Munich
BASF AG
Abstract This work describes the purification, gene cloning and expression of infestin, a thrombin inhibitor from midguts of Triatoma infestans. Infestin is located in the midgut and its purification was performed by anion-exchange and affinity chromatographies. the N-terminal sequence and the sequence of tryptic peptides were determined. Using RT-PCR, total RNA and infestin cDNA information, a DNA fragment was cloned which encodes a multi non-classical Kazal-type serine protease inhibitor. Isolated native infestin has two non-classical Kazal-type domains and shows an apparent molecular mass of 13 kDa, while its gene codes for a protein with four non-classical Kazal-type domains corresponding to an apparent molecular mass of 22 kDa. Two recombinant infestins, r-infestin 1-2 and r-infestin 1-4, were constructed using the vector pVT102U/alpha and expressed in S. cerevisiae. Native and r-infestin 1-2 showed very similar inhibitory activities towards thrombin and trypsin with dissociation constants of 43.5 and 25 pM for thrombin and 2.0 and 3.1 nM for trypsin, respectively. No other serine protease of the blood coagulation cascade was inhibited by the r-infestin 1-2. Surprisingly, r-infestin 1-4 inhibited not only thrombin and trypsin (Ki of 0.8 and 5.2 nM, respectively), but also factor XIIa, factor Xa and plasmin (K-i of 78 pM, 59.2 and 1.1 nM, respectively). (C) 2002 Elsevier B.V. All rights reserved.
Keywords Triatoma infestans
serine protease inhibitor
Kazal-type
blood coagulation
thrombin inhibitor
Language English
Date 2002-09-01
Published in Insect Biochemistry and Molecular Biology. Oxford: Pergamon-Elsevier B.V., v. 32, n. 9, p. 991-997, 2002.
ISSN 0965-1748 (Sherpa/Romeo, impact factor)
Publisher Elsevier B.V.
Extent 991-997
Origin http://dx.doi.org/10.1016/S0965-1748(02)00035-8
Access rights Closed access
Type Article
Web of Science ID WOS:000178361300005
URI http://repositorio.unifesp.br/handle/11600/26967

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