Properties of a purified pore-forming protein (perforin 1) isolated from H-2-restricted cytotoxic T cell granules.

Abstract

Histocompatibility-restricted cytotoxic T lymphocytes produce circular lesions on target cell membranes. The pore-forming protein (PFP or perforin 1) that forms these membrane lesions has been purified from lymphocytes. At 37 degrees C, in the presence of Ca2+, this protein polymerizes into a supramolecular tubular complex of Mr greater than 10(6) that partially resists dissociation by SDS and reducing agents. It incorporates spontaneously into planar lipid bilayers during polymerization to form nonselective ion channels, showing heterogeneous size distribution, the smallest conductance per unit being identified as 400 pS in 0.1 M NaCl. PFP/P1 that had been assembled in lipid vesicles before incorporation into planar bilayer show much larger single channel conductance, ranging from 1 to 6 nS in 0.1 M NaCl, suggesting that PFP/P1 may assume multiple functional sizes in proportion to its state of polymerization. The reconstituted channels are relatively voltage-insensitive, with most channels persisting in the open state for seconds to minutes. Nucleated cells are rapidly depolarized by this protein. The purified protein lyses a variety of tumor cells. Polymerization and functional channel activity are absolutely Ca2+-dependent. The activity of this protein may play a direct role in T lymphocyte-mediated cytolysis.

Calcium-dependent permeabilization of erythrocytes by a perforin-like protein during egress of malaria parasites

“…NATURE COMMUNICATIONS | DOI: 10.1038/ncomms2725 ARTICLE perforin 52 . Deletion of the C-terminal domain of PfPLP1 did not inhibit its membranolytic activity 51 but abolished its Ca 2 þ dependence supporting the fact that the C-terminal region is responsible for conferring its Ca 2 þ dependence similar to human perforin 53 . The C-terminal domain of lymphocyte perforin has been shown to be responsible for Ca 2 þ -dependent membranolytic activity 39,52 .…”

Impaired nonrestricted cytolytic activity in systemic lupus erythematosus

“…TO determine whether a defect in perforin expression could be implicated in the SLE defect, cultured PBMC were stained at various time points for intracellular perforin. Since perforin is associated with cytoplasmic granules (20,23,33,34), the cells were also stained in parallel for expression of the TIA-1 molecule, a 15-kd cytoplasmic granule-associated protein (13). At day 0, discrete antiperforin-and anti-TIA-1-staining populations were readily discernible in both normal and SLE cells (Figure 3).…”

Genomic organization of the mouse pore-forming protein (perforin) gene and localization to chromosome 10. Similarities to and differences from C9.

“…cDNA fragments encoding the 3' terminus ofmouse PFP mRNA were isolated as follows . First-strand cDNAwas synthesized from poly(A)' RNA derived from the cloned cytotoxic T cell line, CTLL-R8 (29), using the oligonucleotide primer 5' GCGGCCGC(T),7 3, exactly as previously described (25). cDNA/RNA molecules were then amplified into double-stranded cDNA by PCR (30), using the above primer and a sense oligonucleotide, whose sequence was derived from the TUT region of the mouse Pfp genomic clone 64.…”