Allotropism in aspartyl-tRNA [transfer ribonucleic acid] synthetase from porcine thyroid

Three forms of aspartyl‐tRNA synthetase, PC‐1, PC‐2 and PC‐3 (based on their order of elution from phosphocellulose columns), were purified from porcine thyroid utilizing fractional pH precipitations and column chromatography. Two forms, PC‐1 and PC‐2, eluted identically from gel filtration columns and migrated the same electrophoretically and in glycerol density gradients. They were estimated to have sedimentation coefficients of 7 S and molecular weights of about 150000. Both PC‐1 and PC‐2 were equally inhibited by KCl. Their elution positions from phosphocellulose were unchanged upon rechromatography, indicating that they were nonequilibrium forms and they were present in post‐mitochondrial supernatants in roughly equal amounts. All three forms were present in the same ratios whether or not phenylmethylsulphonyl fluoride was present in homogenizing buffers. PC‐3 had the characteristics of an aminoacyl‐tRNA synthetase complex. It contained aminoacyl‐tRNA synthetase activity for 16 of 18 amino acids tested and was about 4% low‐molecular‐weight RNA. The complex did not significantly penetrate 5% polyacrylamide gels, was excluded by Sephacryl S‐200 and had a sedimentation coefficient greater than 20 S indicating a particles mass in excess of 500000 daltons. In addition, the PC‐3 fraction contained a ribonuclease capable of degrading both aminoacyl‐tRNA and tRNA. After storage, rechromatography of PC‐3 on phosphocellulose showed that some dissociation of the enzymes had taken place. Aspartyl‐tRNA synthetase activity was found in positions analogous to PC‐1 as well as PC‐3, and ribonuclease activity was found in two positions on the columns, one coincident with PC‐3 and one preceding the position of elution of PC‐1. Dissociation of the complex was also observed using glycerol density gradient centrifugation and by chromatography on Sepharose4B. When present in the PC‐3 complex, the aspartyl‐tRNA synthetase activity and the ribonuclease activity were both stimulated by KC1. However, when dissociated to their free forms, the aspartyl‐tRNA synthetase activity was inhibited by KC1, whereas the ribonuclease remained stimulated. The effects on aspartyl‐tRNA synthetase, appeared to be potassium‐ion‐specific but were more pronounced in the presence of phosphate dianion than in chloride ion. The differential effects of potassium ion on the free and complexed forms of aspartyl‐tRNA synthetase and the prescence of a ribonuclease in the aminoacyl‐tRNA complex that is act