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A 74-kDa δ/B″ subunit was isolated by heparin-Sepharose column chromatography from human erythrocyte protein phosphatase 2 A (PP2A) consisting of a 34-kDa catalytic subunit (α/C) and 63- and 74-kDa regulatory subunits (β/A and δ/B″) in a ratio of 1:1:1. The purified δ/B″ was used as an immunogen in mice, to prepare specific antisera against δ/B″. Immunoblot analyses with the antisera detected an immunoreactive 72-kDa protein in the cytosol from various rat tissues including erythrocytes, brain, lung, testis, adrenal gland, heart, spleen, kidney, and liver. The 72-kDa protein was highly abundant in brain and was distributed evenly in cerebral cortex, cerebellum, and brain stem. The 72-kDa protein was also detected in mitochondria and microsome fractions. An immunoreactive 68-kDa protein was detected mainly in nuclear and microsome fractions. The 72-kDa protein from rat brain cytosol copurified with phosphorylated H2B histone phosphatase activity during successive chromatographies on DEAE-Toyopearl, AH-Sepharose, Sephadex G-150, H1 histone-Toyopearl, TSK DEAE-5PW, protamine-Toyopearl, and TSK G3000SW columns. The purified enzyme migrated as a single protein band on nondenaturing PAGE and as three protein bands of 34, 63, and 72 kDa in a ratio of 1:1:1 on SDS-PAGE. The molecular weight of the enzyme was estimated to be 170,000 from the s_{20, w} value of 7.2 ±0.3 S and the Stokes radius of 5.5 ±0.1 nm. The rat brain enzyme was classified as PP2A, based on the following properties; (1) an IC₅₀ for okadaic acid of 10⁻⁹ M; (2) its preferential dephosphorylation of the α subunit of phosphorylase kinase; (3) its insensitivity to protein inhibitor 2; and (4) its heterotrimeric subunit structure. The K_m value and the molecular activity of the enzyme for phosphorylated H2B histone were 72.3 ± 0.3 μM. and 192±2mol P_i released/min/mol enzyme, respectively, and were comparable to those of human erythrocyte PP2A (α₁β₁/ CAB″). The 72-kDa subunit in the purified rat brain PP2A was phosphorylated in vitro by cAMP-dependent protein kinase.

Aggregates cause a fatal problem in the structural analysis of a biomacromolecule in solution using small-angle X-ray or neutron scattering (SAS): they deteriorate the scattering profile of the target molecule and lead to an incorrect structure. Recently, an integrated method of analytical ultracentrifugation (AUC) and SAS, abbreviated AUC–SAS, was developed as a new approach to overcome this problem. However, the original version of AUC–SAS does not offer a correct scattering profile of the target molecule when the weight fraction of aggregates is higher than ca 10%. In this study, the obstacle point in the original AUC–SAS approach is identified. The improved AUC–SAS method is then applicable to a solution with a relatively larger weight fraction of aggregates (≤20%).

Plasmodium falciparum parasitophorous vacuolar protein 1 (PfPV1), a protein unique to malaria parasites, is localized in the parasitophorous vacuolar (PV) and is essential for parasite growth. Previous studies suggested that PfPV1 cooperates with the Plasmodium translocon of exported proteins (PTEX) complex to export various proteins from the PV. However, the structure and function of PfPV1 have not been determined in detail. In this study, we undertook the expression, purification, and characterization of PfPV1. The tetramer appears to be the structural unit of PfPV1. The activity of PfPV1 appears to be similar to that of molecular chaperones, and it may interact with various proteins. PfPV1 could substitute CtHsp40 in the CtHsp104, CtHsp70, and CtHsp40 protein disaggregation systems. Based on these results, we propose a model in which PfPV1 captures various PV proteins and delivers them to PTEX through a specific interaction with HSP101.