X. TMV STUDIES IN GENETIC CODING 485 



the amino acid composition of the protein. It should be noted that the 

 protein hicks histidine, cystine, and methionine and has but one cysteine. 



The sequential analysis was initiated at the Virus Laboratory at 

 Berkeley in 1952. As usual in the protein field, the structural study 

 began with end group analyses, and in particular with the finding that 

 carboxypeptidase liberated about 2000 threonine residues per virus 

 particle (Harris and Knight, 1952, 1955). This finding served as the 

 first clear indication that the minimal molecular weight of the virus 

 protein represented the subunit or peptide chain weight, a fact which 

 was later supported by physical methods. The structure in the C-terminal 

 region was shown by partial hydrazinolysis to be -Pro-Ala-Thr (Niu and 

 Fraenkel-Conrat, 1955a) and the terminal hexapeptide, -Thr-Ser-Gly- 

 Pro-Ala-Thr, was detected in chymotryptic digests soon thereafter (Niu 

 and Fraenkel-Conrat, 1955b). 



The general methods for amino terminal analysis either failed to 

 yield any knowledge or yielded incorrect information (Braunitzer, 

 1956). The difficulty was explained when Narita (1958) found that an 

 acetyl group blocked the amino-terminal end of the protein. This finding 

 leads to interesting questions concerning the mechanism and the timing 

 of the acetylation of the chain in relation to its biosynthesis and con- 

 cerning the mode by which this feature is controlled by genetic infoiTna- 

 tion. This is a problem of general significance since terminal acetyl 

 groups have since been found to occur not only in all plant viruses 

 investigated, but also in several other proteins, e.g., ovalbumin, cyto- 

 chrome c, and certain hormones. 



Starting from identification of the end groups, the results of a 

 systematic study of the amino acid sequence of the protein were reported 

 from the Virus Laboratory in Berkeley in a series of papers (Gish et al., 

 1958; Ramachandran and Gish, 1959; Gish, 1959, 1960, 1961; Tsugita, 

 1960, 1962a,b; Tsugita et al., 1960) . An independent study by Anderer and 

 co-workers at the Max-Planck-Listitut in Tubingen began to yield valid 

 data in 1960 (Anderer et al., 1960a). The first step in the complete 

 analysis is the splitting of the protein into definite peptides and the 

 separation of these peptides from one another. For this purpose trypsin 

 was the main tool employed by both groups, since other enzymes lack 

 the necessary specificity. Countercurrent distiibution methods were 

 selected for separation of the digested peptides at our laboratory, and 

 more recently basic ion exchange column chromatography (Dowex 1) 

 has been employed. Further purification was necessaiy to obtain most 

 peptides in pure form. The structural studies on some of the individual 

 peptides with as many as 41 residues are rather complicated but almost 

 complete agreement has been reached on the structures of all but this 



