The polypeptides of rat liver nuclear envelope: I. examination of nuclear pore complex polypeptides by solid-state lactoperoxidase labelling

Abstract

Purified nuclei retaining a high degree of ultrastructural integrity were isolated by conventional centrifugation techniques. The cytoplasmic surface of these nuclei was iodinated using lactoperoxidase immobilized onto giant Sepharose beads; thus the outer nuclear membrane and the cytoplasmic surface of nuclear pore complexes were selectively labelled. Pore complexes in association with a fibrous lamina were isolated from these nuclei by removal of the nucleoplasm and extraction with Triton X-100. The chemical composition of the pore-lamina fraction was 93.6% protein, 6% RNA, 0.4% phospholipid. The labelling suggests that major polypeptides N1 (70 000) and N2 (67 000) and more than 10 other more minor polypeptides, ranging from 33 000 to 200 000 mol. wt, as being components of the nuclear pore complex. Polypeptide N3 (58 000) is shown to be present only on the nucleoplasmic face of nuclear envelopes, probably in the fibrous lamina.

Photoaffinity labeling of the major nucleoside triphosphatase of rat liver nuclear envelope

“…(respectively) in our hands. Proteins of the other approximate molecular weights are thought to be associated with the nuclear pore complexes (Richardson & Maddy, 1980a), and thus, these components may also have been labeled by aATP, which was lured to the microenvironment of the pore complex by charge attraction. A nonspecific labeling (particularly of the 58-kDa band) may also in part reflect the fact that the NE NTPase is a relatively low-affinity site.…”