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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Higher-order interactions are important for protein folding and assembly. We introduce the concept of interhelical three-body interactions as derived from Delaunay triangulation and alpha shapes of protein structures. In addition to glycophorin A, where triplets are strongly correlated with protein stability, we found that tight interhelical triplet interactions exist extensively in other membrane proteins, where many types of triplets occur far more frequently than in soluble proteins. We developed a probabilistic model for estimating the value of membrane helical interaction triplet (MHIT) propensity. Because the number of known structures of membrane proteins is limited, we developed a bootstrap method for determining the 95% confidence intervals of estimated MHIT values. We identified triplets that have high propensity for interhelical interactions and are unique to membrane proteins, e.g. AGF, AGG, GLL, GFF and others. A significant fraction (32%) of triplet types contains triplets that may be involved in interhelical hydrogen bond interactions, suggesting the prevalent and important roles of H-bond in the assembly of TM helices. There are several well-defined spatial conformations for triplet interactions on helices with similar parallel or antiparallel orientations and with similar right-handed or left-handed crossing angles. Often, they contain small residues and correspond to the regions of the closest contact between helices. Sequence motifs such as GG4 and AG4 can be part of the three-body interactions that have similar conformations, which in turn can be part of a higher-order cooperative four residue spatial motif observed in helical pairs from different proteins. In many cases, spatial motifs such as serine zipper and polar clamp are part of triplet interactions. On the basis of the analysis of the archaeal rhodopsin family of proteins, tightly packed triplet interactions can be achieved with several different choices of amino acid residues.
J Mol Biol 2003 Mar 14
PMID:Higher-order interhelical spatial interactions in membrane proteins. 1261 23

Congenital adrenal hyperplasia (CAH) is a common autosomal recessive disorder mainly caused by defects in the steroid 21-hydroxylase (CYP21) gene. A 9.3-kb fragment generated by NdeI and AseI digestion by Southern blot analysis indicated that a consequence of deletion of the C4-CYP21 repeat module was the production of a distinct chimeric CYP21P/CYP21 molecule. In the present study, we report a novel CYP21 genotype in two CAH families in which the gene appeared as 9.4- and 3.3-kb fragments by TaqI digestion, rather than as a chimeric gene. From the analysis of PCR amplification patterns and DNA sequencing, we found that there was a duplication of 111 bases from codons 21 to 57 inserted at codon 58 in exon 1 of the CYP21 gene. In addition, codon 21 in the repeated sequence changed from TGG to AGG. Furthermore, this novel CYP21 gene present in both CAH families showed no mutations at IVS2-12A/C>G, 707-714delGAGACTAC, and P30L. Interestingly, the 5' end region of these two CYP21 genes showed the sequence of the CYP21P gene at nucleotides (nt) -103, -110, -123, and thereafter. Our data suggest that these two CYP21 genes are caused by deletion of the CYP21P, XA, RP2, and C4B genes. Possibly, the additional 111-base duplicated coding sequence may be generated by multiple intergenic recombinations, while there seems to be no relationship with deletion of the CYP21P-C4B regions.
Mol Genet Metab 2003 Jul
PMID:Duplication of 111 bases in exon 1 of the CYP21 gene is combined with deletion of CYP21P-C4B genes in steroid 21-hydroxylase deficiency. 1285 27

To analyse the mechanism by which rare codons near the initiation codon inhibit cell growth and protein synthesis, we used the bacteriophage lambda int gene or early codon substitution derivatives. The lambda int gene has a high frequency of rare ATA, AGA and AGG codons; two of them (AGA AGG) located at positions 3 and 4 of the int open reading frame (ORF). Escherichia coli pth (rap) cells, which are defective in peptidyl-tRNA hydrolase (Pth) activity, are more susceptible to the inhibitory effects of int expression as compared with wild-type cells. Cell growth and Int protein synthesis were enhanced by overexpression of Pth and tRNAArg4 cognate to AGG and AGA but not of tRNAIle2a specific for ATA. The increase of Int protein synthesis also takes place when the rare arginine codons AGA and AGG at positions 3 and 4 are changed to common arginine CGT or lysine AAA codons but not to rare isoleucine ATA codons. In addition, overexpression of int in Pth defective cells provokes accumulation of peptidyl-tRNAArg4 in the soluble fraction. Therefore, cell growth and Int synthesis inhibition may be due to ribosome stalling and premature release of peptidyl-tRNAArg4 from the ribosome at the rare arginine codons of the first tandem, which leads to cell starvation for the specific tRNA.
Mol Microbiol 2003 Aug
PMID:The pair of arginine codons AGA AGG close to the initiation codon of the lambda int gene inhibits cell growth and protein synthesis by accumulating peptidyl-tRNAArg4. 1289 27

To elucidate whether the M6p/Igf2 receptor (M6p/Igf2r) gene might be involved in exogenous and endogenous liver carcinogenesis, we investigated its alteration in hepatocellular carcinomas (HCCs) induced by N-nitrosodiethylamine (DEN) and by a choline-deficient L-amino acid-defined (CDAA) diet in rats. Male F344 rats, 6 wk old, received a single intraperitoneal (i.p.) injection of DEN at a dose of 10 mg/kg body weight, followed by combined treatment with partial hepatectomy and colchicine to induce cell cycle disturbance, and a selection procedure regimen, HCCs being obtained after 42 wk. With continuous CDAA diet feeding, tumors were sampled after 75 wk. Total RNA was extracted from individual HCCs for assessment of mutations within exons 27, 28, 31, 33, and 34, and aberrant transcript of the M6p/Igf2r gene by reverse transcription (RT)-polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) and RT-PCR analyses, respectively. Mutations were detected in three of 15 HCCs (20%) induced by the CDAA diet, a TTT to TTG (Phe to Leu) transversion at codon 1516 and two AAG to AGG (Lys to Arg) transitions at codon 1620, but in none of those caused by DEN. Aberrant transcripts were found in seven of 15 HCCs after DEN treatment (46.7%) and in two of 15 HCCs induced by the CDAA diet (13.3%). These results suggest that alterations of the M6p/Igf2r gene may be involved in both exogenous and endogenous liver carcinogenesis with the different patterns and frequencies.
Mol Carcinog 2004 Apr
PMID:Alterations of the M6p/Igf2 receptor gene in hepatocellular carcinomas induced by N-nitrosodiethylamine and a choline-deficient L-amino acid-defined diet in rats. 1505 72

The marRAB multiple antibiotic resistance operon of Escherichia coli is autorepressed by MarR. MarR binds to two palindromic sequences in vitro: site I lies between and overlaps the -35 and -10 hexamers for RNA polymerase binding; site II lies between the transcription start site and the GTG initiation codon of marR. To assess the importance of these sites in vivo, the effects of mutant sites on transcription were analysed using fusions to lacZ in the presence and absence of wild-type MarR. When both sites were wild type, transcription in the derepressed marR-deleted strain was 19-fold that of the wild-type strain; when only site I or site II was wild type, this ratio was reduced to 4.3- and 2.6-fold, respectively, showing that full repression requires both sites, but some repression can occur at one site independently of the other. Translational fusions of the wild-type promoter to lacZ demonstrated that marR translation proceeds at only 4.5% of the transcription rate. Analysis of translational fusions with mutant leader sequences demonstrated that the principal reason for inefficient translation is a weak Shine-Dalgarno (SD) sequence, AGG(G). Although the SD sequence is located within the potential stem-loop structure of site II, no evidence for occlusion of the SD sequence was found in the wild-type strain. However, a single basepair mutation that strengthens the stem-loop structure drastically reduced the translational efficiency. Substitution of ATG for GTG as the initiation codon increased translational efficiency by 50%. Increasing the 5 bp spacing between the SD sequence and the GTG codon by one to four bases reduced the translational efficiency by 50-75%. Inefficient translation of marR may help to sensitize the cell to environmental signals.
Mol Microbiol 2004 Jul
PMID:Transcriptional and translational regulation of the marRAB multiple antibiotic resistance operon in Escherichia coli. 1522 13

For Matthiola incana (Brassicaceae), used as a model system to study biochemical and genetical aspects of anthocyanin biosynthesis, several nearly isogenic colored wild type lines and white-flowering mutant lines are available, each with a specific defect in the genes responsible for anthocyanin production (genes e, f, and g). For gene f supposed to code for chalcone synthase (CHS; EC 2.3.1.74), the key enzyme of the flavonoid/anthocyanin biosynthesis pathway belonging to the group of type III polyketide synthases (PKS), the wild type genomic sequence of M. incana line 04 was determined in comparison to the white-flowering CHS mutant line 18. The type of mutation in the chs gene was characterized as a single nucleotide substitution in a triplet AGG coding for an evolutionary conserved arginine into AGT coding for serine (R72S). Northern blots and RT-PCR demonstrated that the mutated gene is expressed in flower petals. Heterologous expression of the wild type and mutated CHS cDNA in E. Scherichia coli, verified by Western blotting and enzyme assays with various starter molecules, revealed that the mutant protein had no detectable activity, indicating that the strictly conserved arginine residue is essential for the enzymatic reaction. This mutation, which previously was not detected by mutagenic screening, is discussed in the light of structural and functional information on alfalfa CHS and related type III PKS enzymes.
Plant Mol Biol 2004 May
PMID:Characterization and structural features of a chalcone synthase mutation in a white-flowering line of Matthiola incana R. Br. (Brassicaceae). 1560 92

Complete mitochondrial (mt) DNA sequences of two lancelets, Epigonichthys maldivensis and E. lucayanus, were compared with those of two Branchiostoma lancelets and several deuterostomes previously surveyed. The mt-gene order of E. lucayanus was quite different from that of E. maldivensis, the latter being identical to the two Branchiostoma species. A remarkable genomic change in E. lucayanus mtDNA was an inversion, indicating the possibility of recombination of the mt-genome. Gene rearrangements, probably attributable to tandem genome duplications and subsequent random deletions, were observed in two parts. Short major unassignable sequences of the examined lancelets were regarded as a part of putative regulative elements, judging from some sequence similarity to the conserved sequence block (CSB) in mammalian mtDNA. The considerable mt-genome reorganization in E. lucayanus seemed to have affected the nucleotide substitution pattern, suggested by base composition analyses. The present analysis also suggested that AGR codons in lancelet mtDNA were likely to correspond to serine residue, rather than glycine. Furthermore, the AGG codon, so far reputed to be unassignable in lancelet mtDNA, was found twice in E. maldivensis, indicating the availability of all four AGN codons in some lancelets. This finding lends support to an alternative hypothesis regarding the evolutionary history of AGR-codon assignment in extant chordates, rather than that previously proposed. A molecular phylogenetic tree of the Epigonichthys and Branchiostoma species based on DNA sequences of the 13 mt-protein genes doubted the monophyly of the former genus, unlike the prevailing classification based on their different gonadal arrangements.
J Mol Evol 2005 Apr
PMID:Evolution of the mitochondrial genome in cephalochordata as inferred from complete nucleotide sequences from two epigonichthys species. 1588 87

Translating ribosomes can pass through a stretch of messenger RNA without translating and resume protein chain elongation after the bypassed region. We previously investigated the stimulation of bypassing when the codon in the ribosome [corrected] A-site called for an aminoacyl-tRNA species in short supply. Here, we investigate bypassing in unstarved, growing cells. A collection of lacZ bypass reporters was constructed with nearly all the sense codons as the "takeoff site", each with its matched landing site 16 nucleotides downstream in the beta-galactosidase reading frame. Beta-galactosidase [corrected] synthesis in unstarved cells carrying these reporters was found to vary over a large range. The takeoff sites UUU and AGG yielded unusually high enzyme activities, sufficient for protein sequence analysis; in these cases, sequencing (by Edman degradation or by mass spectrometry) confirmed that the synthesis of lacZ protein occurred through the 16 nt bypass from takeoff to landing site. Thus, bypassing occurs spontaneously under normal conditions, and is not limited to the pathology of amino acid starvation. Indirect evidence suggests that most of the lower enzyme activities of the rest of the collection also reflects bypassing. Another collection of reporters was made with [corrected] various triplets in the A-site [corrected] the codon immediately following a UUC [corrected] takeoff triplet. Spontaneous bypassing in representatives of this collection varied roughly inversely with the abundance of the tRNA encoded at the A-site. For two A-site codons tested, introduction of additional copies of the relevant tRNA gene on a second plasmid reduced spontaneous bypassing. We conclude that any pause with the A-site empty stimulates bypassing. From the P-site and A-site effects on bypassing, we estimated the average frequency of ribosome takeoff; from this, we calculate that the probability that a ribosome will succeed in translating the entire lacZ coding sequence is about 0.73, in agreement with earlier, independent estimates.
J Mol Biol 2005 Jun 03
PMID:Spontaneous ribosome bypassing in growing cells. 1589 Jan 94

Programmed translational frameshifting is a ubiquitous but rare mechanism of gene expression in which mRNA sequences cause the translational machinery to shift reading frames with extreme efficiency, up to at least 50%. The mRNA sequences responsible are deceptively simple; the sequence CUU-AGG-C causes about 40% frameshifting when inserted into an mRNA in the yeast Saccharomyces cerevisiae. The high efficiency of this site depends on a set of S. cerevisiae tRNA isoacceptors that perturb the mechanism of translation to cause the programmed translational error. The simplicity of the system might suggest that it could evolve frequently and perhaps be lost as easily. We have investigated the history of programmed +1 frameshifting in fungi. We find that frameshifting has persisted in two structural genes in budding yeasts, ABP140 and EST3 for about 150 million years. Further, the tRNAs that stimulate the event are equally old. Species that diverged from the lineage earlier both do not employ frameshifting and have a different complement of tRNAs predicted to be inimical to frameshifting. The stability of the coevolution of protein coding genes and tRNAs suggests that frameshifting has been selected for during the divergence of these species.
J Mol Evol 2006 Oct
PMID:Evolution of +1 programmed frameshifting signals and frameshift-regulating tRNAs in the order Saccharomycetales. 1683 13

It is generally accepted that the translation rate depends on the availability of cognate aa-tRNAs. In this study it is shown that the key factor that determines translation rate is the competition between near-cognate and cognate aa-tRNAs. The transport mechanism in the cytoplasm is diffusion, thus the competition between cognate, near-cognate and non-cognate aa-tRNAs to bind to the ribosome is a stochastic process. Two competition measures are introduced; C(i) and R(i) (i=1, 64) are quotients of the arrival frequencies of near-cognates vs. cognates and non-cognates vs. cognates, respectively. Furthermore, the reaction rates of bound cognates differ from those of bound near-cognates. If a near-cognate aa-tRNA binds to the A site of the ribosome, it may be rejected at the anti-codon recognition step or proofreading step or it may be accepted. Regardless of its fate, the near-cognates and non-cognates have caused delays of varying duration to the observed rate of translation. Rate constants have been measured at a temperature of 20 degrees C by (Gromadski, K.B., Rodnina, M.V., 2004. Kinetic determinants of high-fidelity tRNA discrimination on the ribosome. Mol. Cell 13, 191-200). These rate constants have been re-evaluated at 37 degrees C, using experimental data at 24.5 degrees C and 37 degrees C (Varenne, S., et al., 1984. Translation in a non-uniform process: effect of tRNA availability on the rate of elongation of nascent polypeptide chains. J. Mol. Biol. 180, 549-576). The key results of the study are: (i) the average time (at 37 degrees C) to add an amino acid, as defined by the ith codon, to the nascent peptide chain is: tau(i)=9.06+1.445x[10.48C(i)+0.5R(i)] (in ms); (ii) the misreading frequency is directly proportional to the near-cognate competition, E(i)=0.0009C(i); (iii) the competition from near-cognates, and not the availability of cognate aa-tRNAs, is the most important factor that determines the translation rate - the four codons with highest near-cognate competition (in the case of E. coli) are [GCC]>[CGG]>[AGG]>[GGA], which overlap only partially with the rarest codons: [AGG]<[CCA]<[GCC]<[CAC]; (iv) based on the kinetic rates at 37 degrees C, the average time to insert a cognate amino acid is 9.06ms and the average delay to process a near-cognate aa-tRNA is 10.45ms and (vii) the model also provides estimates of the vacancy times of the A site of the ribosome - an important factor in frameshifting.
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PMID:Ribosome kinetics and aa-tRNA competition determine rate and fidelity of peptide synthesis. 1789 86


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