By mild treatment of tRNA;:& with acid (pH 2.9, 37", 3-4 hours) the base Y' , which according to RajBhandary, Khorana et al. is located right next to the anticodon, can be excised without breaking the chain. Y+ and the acid treated tRNAPhe (= tRNA&E) were characterized and the conversion of tRNAPhe to tRNAP,h,9 was investigated in detail.1. Yf is a rather hydrophobic compound of unknown structure. Y+ and ribose were found when the nucleoside Y was treated with acid. Some physical and chemical properties of Yf are reported. Yf can be isolated easily in pure form from oligonucleotides, tRNA:,h,e,,, or even unfractionated tRNAyeast (not tRNA,,l() by mild acid treatment and subsequent extraction into chloroform.2. tRNAgEl and tRNAPhe have very similar molecular sizes and shapes since their Sephadex elution profiles were overlapping. The facile separation of tRNAPhe and tRNA&El by countercurrent distribution, on columns of methylated albumin on Kieselgur or benzoylated DEAEcellulose is due, a t least in part, to the removal of a hydrophobic residue from an exposed region of the tRNAPhe.3. tRNA&g can be charged with Phe to the same extent as tRNAPhe. Since also the Michaelis constant of phenylalanyl tRNA synthetase is the same for both tRNAs (1.5 x lo-' M) and the maximal velocity of charging is decreased by only one third in tRNAGE, it can be concluded, that Y and probably the whole anticodon loop are not essential parts of the synthetase recognition region of tRNAPhe in homologous charging reactions. 4.It was found that tRNAgE and tRNAPhe from which the -CpCpA endgroups had been removed do not differ in their acceptor capacities for CTP and ATP.5 . I n the ribosome systems, Phe-tRNAP,h,", does not bind to poly (U) or other polynucleotides and does not transfer its amino acid onto growing peptide chains. The complete loss of codon recognition is attributed largely to a change in the conformation of the anticodon loop. The ultraviolet spectra of an Y-containing hexanucleotide and of its acid conversion products are in agreement with the notion of a strong interaction between Y+ and its neighbouring bases, as are the rate differences in the liberation of Yf from Y 111.5 times faster] and the hexanucleotide, respectively.6. The rate of tRNAEz formation, as measured by methylated albumin on Kieselgur-column chromatography and binding assays, is the same as the rate of Y+-liberation from the hexanucleotide. The acid induced conversion of tRNAPhe to tRNAP,",", is a completely selective reaction affecting Y as the only nucleoside of tRNAPhe.

In this report 118 mouse V O genes are described which, together with the 22 V O genes reported previously (T. Kirschbaum et al., Eur. J. Immunol. 1998. 28: 1458-1466 amount to 140 genes that had been cloned and sequenced in our laboratory. For 73 of them cDNAs are known, i. e. they have to be considered functional genes, although 10 genes of this group have 1-bp deviations from the canonical promoter, splice site or heptanucleotide recombination signal sequences. Twenty V O genes have been defined as only potentially functional since they do not contain any defect, but no cDNAs have been found (yet) for them. Of the 140 V O genes 47 are pseudogenes. There are indications that two to five V O genes or pseudogenes exist in the O locus which we have not yet been able to clone. The 140 V O genes and pseudogenes were assigned to 18 gene families, 4 of them being one-member families. This differs from previous enumerations of the families only by the combination of the V O 9 and V O 10 families and by the addition of the V O dv gene as a new separate family. Sequence identity usually was 80 % or above within the gene families and 55-80 % between genes of different families. Many of the mouse V O gene families show significant homologies to the human ones, indicating that in evolution V O gene diversification predated the divergence of the primate and rodent clades.

Conformational transitions of the anticodon branch, the acceptor branch and the tertiary structure of yeast tRNAPhe were investigated using tRNAPhe fragments. Thermodyn:bmic and kinetic methods were employed using the optical absorbance of the fragments and the fluorescence of the Y-base. Related results of partial nuclease digestions on tRNAPhe fragments are also reported .1. Hypochromicity, reaction enthalpy and rate constants of helix-coil transitions of the anticodon branch agree with the values expected from the cloverleaf model. The dissociation of the branch is accompanied by a quenching of the fluorescence of the Y-base.2. I n some isolated fragments the anticodon branch is more stable than in the intart tRNA. The destabilization in the intact tRNA is attributed to electrostatic effects.3. The anticodon half Phe 21-57, a t low concentrations and temperatures, assumes a structure which is different from a segment of the cloverleaf model. A transition from this structure to the one of a segment of the cloverleaf is observed with increasing temperature. At higher fragment concentrations and in the presence of Mg2+ dimers are formed. 4.Bimolecular recombinations are observed between fragments Phe 1-18 and l'he 21-76 reconstituting the acceptor and dihydrouridine stems, and between Phe 1-18 and Phe 38-76 where only the base pairs of the acceptor stem are newly formed. The mechanism of the recon ibination and the relationship of the fragment combination to the intact tRNA are discussed.5. The thermodynamic properties of the fragment Phe 1-69/70 are very similar to those of the intact tRNA. It can be concluded that the tertiary structure is essentially unchanged anti that the terminal nucleotides 70-76 are not directly involved in its formation. 6. The thermal denaturation process of intact tRNAPhe can now be interpreted a& follows: a t first the tertiary structure is converted to a cloverleaf-like structure ; this process is followed by the melting of the acceptor and anticodon stems ; finally the ribosylthymine and dihyd rouridine stems dissociate independently.Yeast tRNAPhe has been the object of many physico-chemical investigations . The conformational changes with temperature areAbbreviations. The fragments of tRNAPhe are designated by their terminal nucleosides (see also Fig.1). I n addition the following names are used: Phe 1-36, pG-half; Phe 38-76, CCA-half; Phe 21-57, anticodon half; Phe 1-69/70, a mixture of SOo/, Phe 1-69 and 20°/,, Phe 1-70. A,,, unit, the quantity of material contained in 1 ml of a solution which has an absorbance of I a t 260 nm, when measured in an 1-em path length cell.Enzymes. T1-RNAase or ribonucleate guanidine nucleotido-2'-transferase (EC 2.7.7.26) ; N1-RNAase rather complex [5,6]. I n order to resolve the complexit y of the melting process of the intact tRNA, thermodynamic and kinetic studies of tRNAPhe half molecules have been carried out [5]. The cloverleaf model was the basis of the interpretation; the number of base pairs involved, the cooperativity and the nucleation ...

Only 14 of the 25 V kappa genes and pseudogenes had been found before as parts of the L regions. The cloning and linking described in the accompanying report allowed us now to assign to Lp or Ld some V kappa genes which had been found before on scattered clones. In addition the sequences of several still unknown genes are reported here, thus completing the publication of the V kappa genes of the kappa locus as far as they are potentially functional or have only one or two 1-bp defects. Of the V kappa genes of the kappa locus, 32 are potentially functional, 16 have minor defects, 3 have both potentially functional and slightly defective alleles and 25 are pseudogenes which amounts to a repertoire of 76 V kappa-related gene sequences. The V kappa genes of the L regions are, within the subgroups, particularly similar to each other, which is in part due to common evolutionary origins and in part caused by gene conversion-like events. One donor-acceptor pair could be clearly identified, since converted and not-converted alleles of the acceptor gene were found. In other cases the duplicates of the converted genes served as non-converted controls.

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