Low molecular weight squash trypsin inhibitors from Sechium edule seeds.
Nine chromatographic components containing trypsin inhibitor activity were isolated from Sechium edule seeds by acetone fractionation, gel filtration, affinity chromatography and RP-HPLC in an overall yield of 46% of activity and 0.05% of protein. The components obtained with highest yield of total activity and highest specific activity were sequenced by Edman degradation and their molecular masses determined by mass spectrometry. The inhibitors contained 31, 32 and 27 residues per molecule and their sequences were: SETI-IIa, EDRKCPKILMRCKRDSDCLAKCTCQESGYCG; SETI-IIb, EEDRKCPKILMRCKRDSDCLAKCTCQESGYCG and SETI-V, CPRILMKCKLDTDCFPTCTCRPSGFCG. SETI-IIa and SETI-IIb, which differed by an amino-terminal E in the IIb form, were not separable under the conditions employed. The sequences are consistent with consensus sequences obtained from 37 other inhibitors: CPriI1meCk_DSDCla_C_C_G_CG, where capital letters are invariant amino acid residues and lower case letters are the most preserved in this position. SETI-II and SETI-V form complexes with trypsin with a 1:1 stoichiometry and have dissociation constants of 5.4x10(-11)M and 1.1x10(-9)M, respectively.
Laure
Hélen J
HJ
Departamento de Bioquímica, Area de Biologia Molecular, Universidade Federal de São Paulo, Escola Paulista de Medicina, São Paulo, SP, Brazil.
Faça
Vítor M
VM
Izumi
Clarice
C
Padovan
Júlio C
JC
Greene
Lewis J
LJ
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D028464:Cucurbita / Q000737:chemistry*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010940:Plant Proteins / Q000737:chemistry* / Q000302:isolation & purification; D054789:Ribosome Inactivating Proteins, Type 1; D012639:Seeds / Q000737:chemistry*; D014357:Trypsin / Q000737:chemistry / Q000378:metabolism; D014361:Trypsin Inhibitors / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology*
D010940:Plant Proteins; D054789:Ribosome Inactivating Proteins, Type 1; D014361:Trypsin Inhibitors; C114059:sechiumin protein, Sechium edule; D014357:Trypsin