title: Cloning, sequencing, expression, and characterization of thermostability of oligopeptidase B from Serratia proteamaculans, a novel psychrophilic protease.
abstract: Protease from Serratia proteamaculans (PSP) is the first known psychrophilic oligopeptidase B. The gene of S. proteamaculans 94 oligopeptidase B was cloned, sequenced and expressed in Escherichia coli. The unfolding of PSP molecule following heat treatment at 37°C by measuring fluorescence spectra was examined in parallel with the residual activity determination. The effect of PSP thermostabilization by glycerol at 37-50 °С was revealed. Calcium ions and buffer solution of low molarity cause the opposite effect - the acceleration of PSP inactivation at 37°C. The thermal stability of PSP molecule in the presence of 0-100mM CaCl2 was also investigated by means of high-sensitivity differential scanning calorimetry. The artificial reconstruction of the natural complex PSP-chaperonin from S. рroteamaculans was carried out: the stable complex (1:1) of chaperonin E. сoli GroEL with active recombinant enzyme PSP was obtained. It was shown that complex formation with chaperonin promotes PSP thermostability at 37°C.
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Miklukho-Maklaya St. 16/10, Moscow 117997, Russia. Electronic address: anna.mikhailova@ibch.ru.
authors.forename: Anna G
authors.initials: AG
authors.lastname: Mikhailova
authors.forename: Rafil F
authors.initials: RF
authors.lastname: Khairullin
authors.forename: Ilya V
authors.initials: IV
authors.lastname: Demidyuk
authors.forename: Sergey V
authors.initials: SV
authors.lastname: Kostrov
authors.forename: Natalia V
authors.initials: NV
authors.lastname: Grinberg
authors.forename: Tatiana V
authors.initials: TV
authors.lastname: Burova
authors.forename: Valery Ya
authors.initials: VY
authors.lastname: Grinberg
authors.forename: Lev D
authors.initials: LD
authors.lastname: Rumsh
keywords: GroEL; Oligopeptidase B; Psychrophilic enzymes; Serratia proteamaculans; Thermal stability