Purification, inhibitory properties, amino acid sequence and identification of the reactive site of a new serine proteinase inhibitor from oil‐rape (itBrassica napus) seed

FEBS Letters

Bortolotti, Fabrizio²  Ronchi, Severino³  Palmieri, Sandro⁴  Menegatti, Enea²  Ascenzi, Paolo¹  Ceciliani, Fabrizio³ 

[1]Dipartimento di Scienza e Tecnologia del Farmaco, Università di Torino, via Pietro Giuria 9, 10125 Turin, Italy

[2]Dipartimento di Scienze Farmaceutiche, Università di Ferrara, Via Fossato di Mortara 17/19, 44100 Ferrara, Italy

[3]Istituto di Fisiologia Veterinaria e Biochimica, Università di Milano, via Celoria 10, 20133 Milan, Italy

[4]Istituto Sperimentale per Ie Colture Industriali, MAF, Via di Corticella 133, 40129 Bologna, Italy

关键词: Serine proteinase inhibitor;    Amino acid sequence;    Rapeseed;    Brassica napus var. oleifera;    RTI;    rapeseed trypsin inhibitor III;    MTI-2;    low molecular weight mustard trypsin inhibitor;    Bz-l-Arg-PNA;    N-α-benzoyl-l-arginine-p-nitroanilide;    Z-l-Tyr-ONP;    N-α-carbobenzoxy-l-tyrosine-para-nitrophenyl ester;    TFA;    trifluoroacetic acid;    trypsin;    bovine β-trypsin;    chymotrypsin;    bovine α-chymotrypsin;    TPCK-trypsin;    bovine trypsin treated with tosyl-l-phenylalanine chloromethyl ketone;    TLCK-chymotrypsin;    Bovine chymotrypsin trated with N-α-tosyl-l-lysine chloromethyl ketone;   

DOI  :  10.1016/0014-5793(94)80505-9

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

A new serine proteinase inhibitor, rapeseed trypsin inhibitor (RTI), has been isolated from rapeseed (Brassica napus var. oleifera) seed. The protein inhibits the catalytic activity of bovine β-trypsin and bovine α-chymotrypsin with apparent dissociation constants of 3.0 × 10⁻¹⁰ M and 4.1 × 10⁻⁷ M, at pH 8.0 and 21°C, respectively. The stoichiometry of both proteinase-inhibitor complexes is 1:1. The amino acid sequence of RTI consists of 60 amino acid residues, corresponding to an M _r, of about 6.7 kDa. The p₁-p_i, reactive site bond has been tentatively identified at position Arg²⁰-Ile²¹. RTI shows no similarity to other serine proteinase inhibitors except the low molecular weight mustard trypsin inhibitor (MTI-2). RTI and MTI-2 could be members of a new class of plant serine proteinase inhibitors.

【 授权许可】

Unknown