title: Kunitz-type protease inhibitors group B from Solanum palustre.
abstract: Five Kunitz protease inhibitor group B genes were isolated from the genome of the diploid non-tuber-forming potato species Solanum palustre. Three of five new genes share 99% identity to the published KPI-B genes from various cultivated potato accessions, while others exhibit 96% identity. Spls-KPI-B2 and Spls-KPI-B4 proteins contain unique substitutions of the most conserved residues usually involved to trypsin and chymotrypsin-specific binding sites of Kunitz-type protease inhibitor (KPI)-B, respectively. To test the inhibition of trypsin and chymotrypsin by Spls-KPI proteins, five of them were produced in E. coli purified using a Ni-sepharose resin and ion-exchange chromatography. All recombinant Spls-KPI-B inhibited trypsin; K(i) values ranged from 84.8 (Spls-KPI-B4), 345.5 (Spls-KPI-B1), and 1310.6 nM (Spls-KPI-B2) to 3883.5 (Spls-KPI-B5) and 8370 nM (Spls-KPI-B3). In addition, Spls-KPI-B1 and Spls-KPI-B4 inhibited chymotrypsin. These data suggest that regardless of substitutions of key active-center residues both Spls-KPI-B4 and Spls-KPI-B1 are functional trypsin-chymotrypsin inhibitors.
authors.affiliation: Faculty of Biology, M. V. Lomonosov Moscow State University, Vorobjevy Gory, Moscow, Russia.
authors.forename: Anna S
authors.initials: AS
authors.lastname: Speransky
authors.forename: Fabio
authors.initials: F
authors.lastname: Cimaglia
authors.forename: Anastasya A
authors.initials: AA
authors.lastname: Krinitsina
authors.forename: Palmiro
authors.initials: P
authors.lastname: Poltronieri
authors.forename: Pasqua
authors.initials: P
authors.lastname: Fasano
authors.forename: Anna M
authors.initials: AM
authors.lastname: Bogacheva
authors.forename: Tatiana A
authors.initials: TA
authors.lastname: Valueva
authors.forename: Dennis
authors.initials: D
authors.lastname: Halterman
authors.forename: Alexei B
authors.initials: AB
authors.lastname: Shevelev
authors.forename: Angelo
authors.initials: A
authors.lastname: Santino
mesh_terms: D000595:Amino Acid Sequence; D007611:Aprotinin / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D002918:Chymotrypsin / Q000037:antagonists & inhibitors / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D017386:Sequence Homology, Amino Acid; D032322:Solanum / Q000235:genetics / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism; D014361:Trypsin Inhibitors / Q000494:pharmacology
chemical_list: D010940:Plant Proteins; D011994:Recombinant Proteins; D014361:Trypsin Inhibitors; C098973:recombinant Kunitz protease inhibitor; D007611:Aprotinin; D002918:Chymotrypsin; D014357:Trypsin