title: Affinity properties of phosvitin: interaction of phosvitin with serine hydroxymethyl transferase.
abstract: The affinity of phosvitin with serine hydroxymethyl transferase (SHMT), an acidic multi-subunit protein, was evaluated by measurements of enzyme activity, sedimentation velocity, steady-state fluorescence, circular dichroism and kinetic thermal stability. While the presence of phosvitin had no effect on the SHMT activity, the sedimentation coefficient of SHMT increased from 8.7 S to 12.5 S suggesting the formation of a complex at a SHMT:phosvitin molar ratio of 2:1. Based on steady-state fluorescence quenching measurements an association constant of 2.4 +/- 0.2 x 10(5) M-1 at 25 degrees C was obtained for the interaction of phosvitin with SHMT. The temperature dependency of the association constant in the range 15-35 degrees C suggests the involvement of ionic forces in the interaction. The thermal inactivation of SHMT followed first order kinetics. In the presence of phosvitin the rate constant decreased and half time increased. The circular dichroism measurements suggest that phosvitin interaction does not involve pyridoxal phosphate binding domain of the enzyme. Although minor changes in the secondary structure of the enzyme were observed, the environment around aromatic amino acids did not change significantly.
authors.affiliation: Vittal Mallya Scientific Research Foundation, Bangalore, India.
authors.forename: H V
authors.initials: HV
authors.lastname: Lakhey
authors.forename: A G
authors.initials: AG
authors.lastname: Rao
authors.forename: V
authors.initials: V
authors.lastname: Prakash
authors.forename: P R
authors.initials: PR
authors.lastname: Krishnaswamy
authors.forename: H S
authors.initials: HS
authors.lastname: Savithri
authors.forename: N A
authors.initials: NA
authors.lastname: Rao
authors.forename: C S
authors.initials: CS
authors.lastname: Ramadoss
mesh_terms: D005453:Fluorescence; D012696:Glycine Hydroxymethyltransferase / Q000378:metabolism*; D010774:Phosvitin / Q000378:metabolism*; D014461:Ultracentrifugation
chemical_list: D010774:Phosvitin; D012696:Glycine Hydroxymethyltransferase