摘要:

We have applied the SELEX procedure (systematic evolution of ligands by exponential enrichment) to obtain RNA molecules that bind tightly to theEscherichia colitranscription termination factor rho. The starting pool was a population of RNA molecules 77 nucleotides (nt) long, in which was embedded a cassette of 30 nt of randomized sequence. The apparent dissociation constant of this RNA pool for hexameric rho factor was about 1μM. After eight rounds of selection by filter binding, with RNA in either 10‐fold or 40 to 100‐fold excess at each step, the dissociation constant of the selected RNA had dropped by more than 500‐fold to about 1 nM. Analysis of 29 clonal isolates from the population revealed that five hadKDSsubstantially weaker than 10 nM (presumably background carryover), 40% were C‐rich (as might have been predicted from rho's known substrate binding), and 40% had a strikingly preserved potential hairpin, in most cases of 6 base pairs with a 3 nt CAA loop and preceded by a CCCCA consensus. The rhodependenttrp t'terminator region includes a related potential hairpin structure; however, it is energetically unfavorable. The implications of the sequence findings for elucidating both static and dynamic aspects of rho factor recognition and response to its RNA target site are discussed.— Schneider, D., Gold, L., Platt, T. Selective enrichment of RNA species for tight binding toEscherichia colirho factor.FASEB J.7: 201‐207; 1993.

ISSN:0892-6638    

DOI:10.1096/fasebj.7.1.7678562

出版商:Wiley    

年代:1993

数据来源: WILEY

摘要:

RNase MRP is a ribonucleoprotein endoribonuclease that has been shown to cleave mitochondrial primer RNA sequences from a variety of sources. The bulk of RNase MRP activity is found in the nucleus where its function remains unknown. Two different approaches have resulted in predictions of distinct secondary structures for RNase MRP RNA. In order to analyze more definitively the higher‐order structure of RNase MRP RNA, we have conducted a phylogenetic comparison of the available RNase MRP RNA sequences from human, mouse, rat, cow, toad, and yeast. The resulting secondary structure shares features in common with previously described structures for prokaryotic and eukaryotic RNase P RNAs (1) and RNase MRP RNAs (2, 3). In addition, the phylogenetic structure is consistent with available chemical modification data on RNase MRP RNA and with the detailed analysis of the To antigen binding domain located near the 5′ end of the RNase MRP RNA. The structure is not limited to RNase MRP RNAs, but can be expanded to cover both eukaryotic RNase P RNAs and RNase P/MRP RNAs from plants.— Schmitt, M. E., Bennett, J. L., Dairaghi, D. J., Clayton, D. A. Secondary structure of RNase MRP RNA as predicted by phylogenetic comparison.FASEB J.7: 208‐213; 1993.

ISSN:0892-6638    

DOI:10.1096/fasebj.7.1.7678563

出版商:Wiley    

年代:1993

数据来源: WILEY

摘要:

Human immunodeficiency virus (HIV‐1) gene expression is activated by the viral TAT protein that interacts with an RNA sequence, TAR, located at the 5′ end of all viral mRNAs. TAT functions primarily as a transcriptional activator in mammalian cells. However, inXenopusoocytes TAT functions primarily as a translational activator. TAR is an RNA structure comprising a partially base‐paired stem, a tripyrimidine bulge in the upper stem, and an unpaired six‐nucleotide loop. In vitro, TAT binds directly to the bulge with no requirement for the loop. In vivo, however, mutations in the loop abolish TAT activation of transcription and translation, implying a requirement for TAR‐binding cellular factors. We now provide genetic evidence for the presence of two TAR‐specific cellular factors inXenopusoocytes. These factors display independent and mutually exclusive interactions with either the loop or the bulge region of TAR. Furthermore, by using in vivo RNA competition assays we show that the cellular factors regulate the accessibility of the TAT binding site. The fact thatXenopusoocytes contain factors that specifically interact with a human viral RNA sequence might indicate that the TAT/TAR interaction is subverting a conserved pathway in the cell.— Braddock, M., Powell, R., Blanchard, A. D., Kingsman, A. J., and Kingsman, S. M. HIV‐1 TAR RNA binding proteins control TAT activation of translation inXenopus oocytes.FASEB J.7: 214‐222; 1993.

ISSN:0892-6638    

DOI:10.1096/fasebj.7.1.8422967

出版商:Wiley    

年代:1993

数据来源: WILEY

摘要:

Complete small‐subunit rRNA (SSU‐rRNA) coding region sequences were determined for two species of the intestinal parasiteGiardia: G. ardeaeandG. muris, both belonging to the order Diplomonadida, and a free‐living member of this order,Hexamita sp.These sequences were compared to published SSU‐rDNA sequences from a third member of the genusGiardia, G. duodenalis(often calledG. intestinalisorG. lamblia) and various representative organisms from other taxa. Of the threeGiardiasequences analyzed, the SSU‐rRNA fromG. murisis the smallest (1432 bases as compared to 1435 and 1453 forG. ardeaeandG. duodenalis, respectively) and has the lowest G + C content (58.9%). TheHexamitaSSU‐rRNA is the largest in this group, containing 1550 bases. Because the sizes of the SSU‐rRNA are prokaryotic rather than typically eukaryotic, the secondary structures of the SSU‐rRNAs were constructed. These structures show a number of typically eukaryotic signature sequences. Sequence alignments based on constraints imposed by secondary structure were used for construction of a phylogenetic tree for these four taxa. The results show that of the four diplomonads represented, theGiardiaspecies form a distinct group. The other diplomonadHexamitaand the microsporidiumVairimorpha necatrixappear to be distinct fromGiardia.—van Keulen, H., Gutell, R. R., Gates, M. A., Campbell, S. R., Erlandsen, S. L., Jarroll, E. L., Kulda, J., Meyer, E. A. Unique phylogenetic position of Diplomonadida based on the complete small subunit ribosomal RNA sequence ofGiardia ardeae, G. muris, G. duodenalis, andHexamita sp.FASEB J.7: 223‐231; 1993.

ISSN:0892-6638    

DOI:10.1096/fasebj.7.1.8422968

出版商:Wiley    

年代:1993

数据来源: WILEY

摘要:

In order to investigate the genetic diversity of streptomycetes in an acid forested soil sample from Mt. Coot‐tha, Brisbane, Australia, cells were mechanically lysed within the soil matrix and genomic DNA was isolated and purified. 16S ribosomal (r)DNA was amplified by the polymerase chain reaction (PCR) method using one primer conserved for members of the domain Bacteria and a second designed specifically for streptomycetes and related taxa. PCR amplification products were cloned into phage vector M13 mp19 and the diversity of 16S rDNA genes was determined by sequence analysis and oligonucleotide probing of the resultant clone library. Comparison of partial 16S rDNA sequences with published sequences revealed that few sequences originated from streptomycetes. The majority of sequences belonged to members of the alpha subclass of Proteobacteria. Other clones were related to planctomycetes, actinomycetes, or represented novel lines of descent. Bacteria that are customarily isolated from soil of pH 4‐7 such as thiobacilli, bacilli, spore‐ and nonsporeforming actinomycetes, and pseudomonads are represented in the clone library in small numbers or were not detected at all. Parameters influencing the recovery, amplification, quantification, and interpretation of genetic information from natural sites are discussed.— Stackebrandt, E., Liesack, W., Goebel, B. M. Bacterial diversity in a soil sample from a subtropical Australian environment as determined by 16S rDNA analysis.FASEB J.7: 232‐236; 1993.

ISSN:0892-6638    

DOI:10.1096/fasebj.7.1.8422969

出版商:Wiley    

年代:1993

数据来源: WILEY

ISSN:0892-6638    

DOI:10.1096/fasebj.7.1.8422970

出版商:Wiley    

年代:1993

数据来源: WILEY

ISSN:0892-6638    

DOI:10.1096/fasebj.7.1.7678564

出版商:Wiley    

年代:1993

数据来源: WILEY