Human lamin B contains a farnesylated cysteine residue.
We recently showed that HeLa cell lamin B is modified by a mevalonic acid derivative. Here we identified the modified amino acid, determined its mode of linkage to the mevalonic acid derivative, and established the derivative's structure. A cysteine residue is modified because experiments with lamin B that had been biosynthetically labeled with [3H]mevalonic acid or [35S]cysteine and then extensively digested with proteases yielded 3H- or 35S-labeled products that co-chromatographed in five successive systems. A thioether linkage rather than a thioester linkage is involved because the mevalonic acid derivative could be released from the 3H-labeled products in a pentane-extractable form by treatment with Raney nickel but not with methanolic KOH. The derivative is a farnesyl moiety because the Raney nickel-released material was identified as 2,6,10-trimethyl-2,6,10-dodecatriene by a combination of gas chromatography and mass spectrometry. The thioether-modified cysteine residue appears to be located near the carboxyl end of lamin B because treatment of 3H-labeled lamin B with cyanogen bromide yielded a single labeled polypeptide that mapped toward this end of the cDNA-inferred sequence of human lamin B.
Farnsworth
C C
CC
Howard Hughes Medical Institute Laboratory, University of Washington, Seattle 98195.
Wolda
S L
SL
Gelb
M H
MH
Glomset
J A
JA
D000595:Amino Acid Sequence; D002467:Cell Nucleus / Q000032:analysis; D003488:Cyanogen Bromide; D003545:Cysteine / Q000302:isolation & purification / Q000378:metabolism / Q000031:analogs & derivatives; D008401:Gas Chromatography-Mass Spectrometry; D006367:HeLa Cells / Q000032:analysis; D006801:Humans; D007202:Indicators and Reagents; D034921:Lamin Type B; D034882:Lamins; D008798:Mevalonic Acid / Q000378:metabolism; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000096:biosynthesis / Q000302:isolation & purification*; D010447:Peptide Hydrolases; D012689:Sequence Homology, Nucleic Acid; D013462:Sulfur Radioisotopes; D014316:Tritium
D007202:Indicators and Reagents; D034921:Lamin Type B; D034882:Lamins; D009687:Nuclear Proteins; D013462:Sulfur Radioisotopes; D014316:Tritium; C057785:S-farnesylcysteine; D010447:Peptide Hydrolases; D003545:Cysteine; D003488:Cyanogen Bromide; D008798:Mevalonic Acid