IDENTIFICATION OF BACTERIOPHAGE-T4 VIRION PROTEINS BY TRANSVERSE PORE-GRADIENT SODIUM DODECYL SULFATE-POLYACRYLAMIDE GEL-ELECTROPHORESIS AND DUAL AMINO-ACID LABELING

We have developed a horizontal N, N'-methylenebisacrylamide (Bis) acry lamide gradient sodium dodecyl sulfate (SDS) gel system that permits t he evaluation of the purity of individual protein bands in complex mix tures. A Bis gradient gel is poured vertically and, after polymerizati on, reoriented horizontally. a single large sample spanning the top of the gel is then run down at right angles to the gradient. The relativ e mobility of a few proteins varies considerably from the rest, causin g them to merge with and cross other bands as the Bis concentration ch anges. Band splitting revealed that several bands previously thought t o represent a single species are actually comprised of comigrating pro teins, Once the Bis/monomer concentration offering the best separation was identified, we sought a simple method for identifying the genetic origin of bands, since many proteins now migrated in new positions on the gel. We reasoned that if infected cells were simultaneously label ed with [S-35]methionine and [H-3]leucine and the purified virion prot eins analyzed to determine their S-35/H-2 ratio, we could identify mos t proteins by comparing this ratio with one calculated from the T4 DNA sequence, Our expectations were realized, and we here report the sepa ration and identification of all T4 virion proteins. Finally, we comme nt on the incorporation of various changes to the original Laemmli SDS -polyacrylamide gel formulations that have been reported in the litera ture.