期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：1977
卷号：74
期号：7
页码：2830-2834
DOI：10.1073/pnas.74.7.2830
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：To provide direct evidence for the hypothesis that secreted proteins may traverse membranes as growing chains, we labeled spheroplasts of Escherichia coli with a reagent (acetyl[35S]methionyl methylphosphate sulfone) that reacts with amino groups but does not cross the membrane. After fractionation, about 6% of the label in the membrane-polysome fraction was found to be attached to the polysomes. This attachment was via peptidyl-tRNA, as shown by several tests: release of most of the label from purified polysomes at low Mg2+; subsequent loss of about 25,000 daltons on cleavage by dilute alkali; release by puromycin; and release, accompanied by a marked increase in average molecular weight, on peptide chain completion. Moreover, a significant fraction of the completed chains was identified serologically and by molecular weight as a major periplasmic protein, alkaline phosphatase [orthophosphoric-monoester phosphohydrolase (alkaline optimum); EC 3.1.3.1 ]. This work provides direct evidence that: (i) secreted proteins thread through the membrane as growing peptide chains; and (ii) membrane-associated polysomes in bacteria are functionally attached to membrane and not merely trapped on disruption of the cell.
