摘要:

AbstractThe conformation of native double helical DNA is well‐known, but it is possible that small regions occur within native DNA, undetectable by X‐ray diffraction methods, which have different conformations. Model structures are the synthetic deoxypolynucleotides of defined sequence. Under the conditions used, DNA, poly d(A‐T) • poly d(A‐T), and poly d(T‐G) • poly d(C‐A) can all give similar X‐ray diffraction patterns, whereas poly dA • poly dT, poly dI • poly dC, poly dG • poly dC, and poly d(T‐C) • poly d(G‐A) clearly differ from DNA. This led to the tentative hypothesis that those DNA's in which all purines are in one strand and all pyrimidines in the other differ in structure from those (such as native DNA) in which purines and pyrimidines alternate or are irregular. We now find that poly d(I‐C) • poly d(I‐C) does not fit the hypothesis and is a most unusual structure, having seven or eight base pairs per turn. Both molecular model building and circular dichroism studies suggest that it is a left‐handed helix. A number of purified tRNA's have been crystallized. We have obtained, from unfractionated tRNA, crystalline “powder” X‐ray diffraction patterns showing rings and spots to about 20 Å resolution. It is not clear whether cocrystallization has occurred, or whether there is fractional crystallization, though preliminary evidence favors the latter. Determination of the structure of crystalline tRNA has many features in common with protein crystallography, bu

ISSN:0021-9541    

DOI:10.1002/jcp.1040740403

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1969

数据来源: WILEY

摘要:

AbstractMaturation of bacterial viruses requires the formation of mature viral nucleic acid in the form found in progeny virus particles, the “packaging” of this viral nucleic acid within a capsule, and the assembly of “packaged” nucleic acid with the accessory structures necessary for infectivity. Recognition that the replicating viral nucleic acid is frequently in a form distinct from that found in mature particles has, in some instances, led to the postulation and formal description of presently unknown processes that must participate in the synthesis of the mature form. In at least some instances, packaging of the mature nucleic acid is clearly integrated with its synthesis.Examples involving the DNA bacteriophages T4, λ, and ϕ X174 and the RNA bacteriophages are presented to illustrate the

ISSN:0021-9541    

DOI:10.1002/jcp.1040740404

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1969

数据来源: WILEY

摘要:

AbstractThe development of a virus is programmed by a series of negative and positive controls which determine the timing and the segment on either of the two DNA strands (lorr) to be transcribed into specific messenger RNA's. Bacteriophage λ provides one of the most deeply studied systems for following the development of lysogenic viruses. In the lysogenic repressed state, only 2–4% of the λ genome is expressed. Thispc‐cI‐rexregion is transcribed leftward to produce a repressor protein which prevents any further transcription by blocking theoLandoRoperators flanking thecI‐rexoperon (figs. 1, 2). Thisnegativecontrol is relieved by destruction of the repressor, and the result is the “induction” of viral development. The earliest post‐induction or postinfection events are the leftward transcription of thepLoLN region from strandland the rightward transcription mainly of thepRoR‐xsegment from strandr. The N product acts as apositivecontrol, permitting a leftward transcription beyond gene N and a rightward transcription of genescII‐O‐P and also Q. Theint‐xissystem controls the excision of the λ genome, whereas the act of rightward transcription and the products of genes O and P initiate the replication of λ DNA. The product of gene Q, still anotherpositivecontrol, stimulates rightward transcription of the late genes which control the synthesis and assembly of the phage heads and tails as well as cell lysis. Among other types ofnegativecontrol are the possible competition between the two divergent transcriptions originating in regionx, the “antirepressor” effect of thexproduct, and the interference between the two convergent transcriptions which collide in the central b2 region. The majority of controls are based on protein‐DNA interactions and can be modified by mutations. For instance, transcription can be rendered independent of negative repressor control either by constitutive,v, mutations which decrease or abolish the affinity of theooperators for the repressor or by insertion of new promoters–e.g., c17orric‐ on the “downstream” side of the operator. The need for the positive N and Q controls may also be obviated by mutations in the N‐ or Q‐dependent promoter or terminator elements.The specific DNA structure within the controlling sites is not known. However, a remarkable coincidence was observed; namely, the occurrence of pyrimidine‐rich clusters in those segments of the individual DNA strands acting as templates for RNA synthesis. This observation, which pertains to all studied DNA's, including those of phages T2, T3, T4, T5, T6, T7, λ, and ϕ 80, formed the basis for a proposal that implicates pyrimidine‐rich clusters in the initiation, control and/or termination of transcription, and also in the determination of the preferred strand and, consequently, the orientation of transcription. General considerations regarding the possible role of the structural singularities, especially those represented by the pyrimidine clusters, in the bipartite structur

ISSN:0021-9541    

DOI:10.1002/jcp.1040740405

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1969

数据来源: WILEY

摘要:

AbstractT4 bacteriophage infection ofE. coliB cells induces the formation of 4S RNA molecules that specifically hybridize with T4 DNA. The T4 4S RNA extracted from the hybrid was found to contain pseudouridylic acid, suggesting that some of this RNA might have amino acid acceptor function. In order to study the amino acid acceptor capacity of the T4 4S RNA, two procedures were devised. The first one involved the isolation and purification of specific 4S RNA‐DNA hybrids in a manner that avoided the use of RNase and permitted the extraction of biologically active tRNA from the hybrid. It was found that a significant fraction of the T4 4S RNA isolated by this method had amino acid acceptor activity. This was shown by assaying with a mixture of 1514C‐labeled amino acids or with [14C]leucine alone.In the second method, T4N‐acetyl[3H]aminoacyl‐tRNA was prepared in order to stabilize the aminoacyl‐tRNA ester bond. T4N‐acetyl[3H]leucyl‐tRNA was incubated with T4 DNA in the presence of 50% formamide at 30°C. Sephadex G‐200 chromatography revealed that a significant fraction of theN‐acetyl[3H]leucyl‐tRNA hybridized with the T4 DNA. Another procedure involved the hybridization ofN‐acetyl[3H]‐aminoacyl‐tRNA with T4 DNA at 70°C in a citrate buffer in the absence of formamide. The annealing mixture also contained a 20‐fold excess of unchargedE. colitRNA. The hybrid‐containing solution was loaded onto nitrocellulose filters and treated with T1 RNase. At this point, the T4 tRNA was found to contain leucine‐, arginine‐, isoleucine‐,

ISSN:0021-9541    

DOI:10.1002/jcp.1040740406

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1969

数据来源: WILEY

ISSN:0021-9541    

DOI:10.1002/jcp.1040740407

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1969

数据来源: WILEY

摘要:

AbstractInfection ofEscherichia coli: with T4 bacteriophage causes the appearance of a new valyl‐tRNA synthetase activity associated with a molecule that, compared to the host enzyme, exhibits a greater resistance to denaturation by heat or urea, a larger molecular volume, a higher rate of sedimentation in sucrose gradients, a greater net positive charge, and a greater ability to charge yeast tRNA. No evidence has been found for similar changes in synthetase activity for the other amino acids. Appearance of the new activity requires continued protein synthesis and results from a modification of the preexisting host enzyme rather thande novosynthesis of a totally new enzyme. By 20 minutes after infection at 30°C, all of the host enzyme has been converted into the new form. Phage mutants have been isolated that fail to effect a normal conversion. The properties of these mutants suggest that conversion involves the addition, to the host enzyme, of a protein specified by the phage genome. Drastic reduction of phage‐induced activity in one of these mutants does not interfere detectably with phage development in a normal host, suggesting that the presence of the new activity is not essential for normal phage produc

ISSN:0021-9541    

DOI:10.1002/jcp.1040740408

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1969

数据来源: WILEY

摘要:

AbstractPartially purified preparations of two protein factors, one of which is stable at 50°C (FIs) and the other unstable at 50°C (FIu), are required for the GTP‐dependent, mRNA‐directed binding of aminoacyl‐tRNA to ribosomes, as well as for polypeptide synthesis in the presence of a third factor, FII. Both FIsand FIuare required for maximal interaction with GTP to form a GTP•protein complex that subsequently interacts with aminoacyl‐tRNA, but not with deacylated tRNA or withN‐(substituted)‐aminoacyl‐tRNA, to form an aminoacyl‐tRNA•GTP•protein complex. A mixture of FIsand FIualso interacts with GDP to form a GDP•protein complex; however, no subsequent interaction with aminoacyl‐tRNA is observed. In addition to aminoacyl‐tRNA and GTP, Mg2+and NH4+are required for the formation of the aminoacyl‐tRNA•GTP•protein complex. Although both protein factors, FIsand FIu, are required for the formation of this complex, only the heat‐labile protein, FIu, is a component of the complex. Very little dissociation of the GTP moiety of the complex occurs in the presence of Mg2+, and no detectable exchange is observed with GTP, GDP, or Pi. In contrast, appreciable dissociation of the aminoacyl‐tRNA from the GTP•protein occurs even in the presence of Mg2+, and exchange with other aminoacyl‐tRNA's can be readily demonstrated. In the absence of Mg2+, complete dissociation of both the GTP and the aminoacyl‐tRNA from the protein occurs. Evidence has been obtained to demonstrate that the aminoacyl‐tRNA•GTP•protein complex is an intermediate in the GTP‐dependent binding of aminoacyl‐tRNA to ribosomes. The binding of the aminoacyl‐tRNA to the ribosome occurs with the concomitant formation of Piand a GDP • protein complex. Incorporation of the bound aminoacyl‐tRNA into polypepti

ISSN:0021-9541    

DOI:10.1002/jcp.1040740409

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1969

数据来源: WILEY

ISSN:0021-9541    

DOI:10.1002/jcp.1040740410

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1969

数据来源: WILEY

摘要:

AbstractAfter the experimental verification of Crick's adaptor hypothesis for the role of tRNA, it became apparent that one of the most important of the protein‐nucleic acid interactions occurs at the first step in protein synthesis, namely the amino acid activation reaction. It is here that a specific aminoacyl‐tRNA synthetase must select, with high fidelity, a specific tRNA out of a large collection of molecules of similar size, shape, and overall composition. A mistake at this point, either by esterification of the wrong amino acid to the correct tRNA or by selection of the wrong tRNA, will inevitabley result in the insertion of an amino acid at an incorrect position in a growing polypeptide. Although there are known rules that dictate how one nucleic acid can recognize and interact with another nucleic acid, nothing is known regarding the mechanism by which a specific protein can recognize and interact with a specific nucleic acid. In order to gain some insight into the specific recognition between an aminoacyl‐tRNA synthetase and its cognate tRNA, it became necessary to study the specific interaction with highly purified materials, preferably in gram quantities. An effort to do this for both the synthetases and the tRNA's was launched at the Oak Ridge National Laboratory about 6 years ago. Four high‐resolution column chromatographic procedures have been developed in the ORNL Macromolecular Separations Program for the separation and production of highly purified species of tRNA's. An unexpected “spin‐off” from this program is the analytical use of some of these systems to detect qualitative changes in the tRNA profile of cells as a consequence of virus infection, methionine starvation, and other metabolic alterations.Some examples of the heterologous interaction between aminoacyl‐tRNA synthetases of one species with the tRNA's of another species, and some of the inherent dangers in the interpretation of such interactions, are considered.Finally, some speculations are made regarding the possible role of tRN

ISSN:0021-9541    

DOI:10.1002/jcp.1040740411

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1969

数据来源: WILEY

ISSN:0021-9541    

DOI:10.1002/jcp.1040740412

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1969

数据来源: WILEY