The SKS of the KMSKS signature of class I aminoacyl-tRNA synthetases corresponds to the GKT/S sequence characteristic of the ATP-binding site of many proteins

On the basis of protein modification studies and primary structure comparison, we propose that the SKS sequence within the KMSKS signature of the class 1 aminoacyl-tRNA synthetases corresponds to the GKT(or S) sequence considered as a signature of the nucleotide triphosphate-binding site of many proteins.     

Sequence and structural homology among membrane-associated domains of CFTR and certain transporter proteins

A sequence comparison of the two membrane-associated (MA) domains of the cystic fibrosis transmembrane conductance regulator (CFTR), multidrug resistance transporter (MDR), and -factor pheromone export system (STE6) proteins, each of which are believed to contain a total of 12 transmembrane (TM) segments, reveals significant amino acid homology and length conservation in the loop regions that connect individual TM sequences. Similar structural homology is observed between these proteins, hemolysin B (HLYB) and the major histocompatibility-linked peptide transporter, HAM1, the latter two which contain a single MA domain composed of six TM segments. In addition, there are specific sequences that are conserved within the TM segments of the five different membrane proteins. This observation suggests that the folding topologies of the MA domains of MDR, STE6, and CFTR in the plasma membrane are likely to be very similar. The sequence analysis also reveals that there are three characteristic motifs (a pair of aromatic residues, LTLXXXXXXP and GXXL) that are conserved in MDR, STE6, HLYB, HAM1, but not in CFTR. We propose that although CFTR may be evolutionarily related to these other membrane proteins, it belongs to a separate subclass.     

ThechlL (frxC) gene: Phylogenetic distribution in vascular plants and DNA sequence fromPolystichum acrostichoides (Pteridophyta) andSynechococcus sp. 7002 (Cyanobacteria)

We examinedchlL (frxC) gene evolution using several approaches. Sequences from the chloroplast genome of the fernPolystichum acrostichoides and from the cyanobacteriumSynechococcus sp. 7002 were determined and found to be highly conserved. A complete physical map of the fern chloroplast genome and partial maps of other vascular plant taxa show thatchlL is located primarily in the small single copy region as inMarchantia polymorpha. A survey of a wide variety of non-angiospermous vascular plant DNAs shows thatchlL is widely distributed but has been lost in the pteridophytePsilotum and (presumably independently) within the Gnetalean gymnosperms.The namefrxC was originally used to denote a gene encoding a product with probable Fe : S cluster binding activity. This activity was postulated due to the amino acid sequence similarity between this product and the Fe : S-binding nitrogenase iron proteinnifH. Fe : S-binding is a property shared by ferredoxins, which are denoted by the prefix frx. However, this gene does not encode a ferredoxin. It is much larger than any known ferredoxin, it binds its Fe : S cluster between two halves of a homodimer (Fujita & al. 1989,Burke & al. 1993 a, c) instead of within a single subunit, and it lacks the pattern of clustered cysteines present in all ferredoxins (Meyer 1988). Therefore, we use the namechlL to recognize the sequence and functional similarities to the bacterial PChlide reductase subunit,bchL. Similar usage has been adopted for this (Suzuki & Bauer 1992) and other (Choquet & al. 1992,Burke & al. 1993b) PChlide reductase subunits.     

Experimental and computational analysis of the ancestry of an evolutionary young enzyme from histidine biosynthesis

The conservation of fold and chemistry of the enzymes associated with histidine biosynthesis suggests that this pathway evolved prior to the diversification of Bacteria, Archaea, and Eukaryotes. The only exception is the histidinol phosphate phosphatase (HolPase). So far, non-homologous HolPases that possess distinct folds and belong to three different protein superfamilies have been identified in various phylogenetic clades. However, their evolution has remained unknown to date. Here, we analyzed the evolutionary history of the HolPase from γ-Proteobacteria (HisB-N). It has been argued that HisB-N and its closest homologue d -glycero-d -manno-heptose-1,7-bisphosphate 7-phosphatase (GmhB) have emerged from the same promiscuous ancestral phosphatase. GmhB variants catalyze the hydrolysis of the anomeric d -glycero-d -manno-heptose-1,7-bisphosphate (αHBP or βHBP) with a strong preference for one anomer (αGmhB or βGmhB). We found that HisB-N from Escherichia coli shows promiscuous activity for βHBP but not αHBP, while βGmhB from Crassaminicella sp. shows promiscuous activity for HolP. Accordingly, a combined phylogenetic tree of αGmhBs, βGmhBs, and HisB-N sequences revealed that HisB-Ns form a compact subcluster derived from βGmhBs. Ancestral sequence reconstruction and in vitro analysis revealed a promiscuous HolPase activity in the resurrected enzymes prior to functional divergence of the successors. The following increase in catalytic efficiency of the HolP turnover is reflected in the shape and electrostatics of the active site predicted by AlphaFold. An analysis of the phylogenetic tree led to a revised evolutionary model that proposes the horizontal gene transfer of a promiscuous βGmhB from δ- to γ-Proteobacteria where it evolved to the modern HisB-N.     

Drosophila acidic ribosomal protein rpA2: Sequence and characterization

A cDNA encoding the Drosophila melanogaster acidic ribosomal protein rpA2 was cloned and sequenced. rpA2 is homologous to the Artemia salina acidic ribosomal protein eL12′. In situ hybridization to salivary gland polytene chromosomes localizes the rpA2 gene to band 21C. It is a single copy gene, with an mRNA of 0.8 kb. Two-dimensional gel electrophoresis of Drosophila ribosomal proteins followed by immuno-blotting showed that the rpA2 protein has an apparent relative mobility in SDS of 17 kD and an isoelectric point less than pH 5.0. Although the Drosophila gene rp21C may be the same as rpA2, the reported sequences differ. Comparisons of the aligned nucleotide sequences coding for the acidic ribosomal proteins rpA1 and rpA2 of Drosophila with those of other eukaryotes support the view of two separate, though closely related, groups of acidic proteins. Comparison with the Artemia homologues suggests that nucleotide identity may have been conserved by some constraint that acts in addition to the requirement for substantial similarity of amino acid sequences. © 1993 Wiley-Liss, Inc.     

Molecular cytological evidence for gradual telomere synthesis at the broken chromosome ends in wheat

Telomere formation of the normal and broken chromosomes of common wheat,Triticum aestivum, was investigated byin situ hybridization using the biotin-labeled probe of telomere repetitive sequences (pAtT4) ofArabidopsis thaliana with subsequent amplification by an antibody. After double and triple amplification, prominent signals appeared at all the telomeric regions of the normal chromosomes. Prominent signals also emerged at the broken ends of the telocentric and deletion chromosomes that had passed through more than one generation since the appearance. However, broken ends that had passed through only the stages of gametogenesis, fertilization, embryogenesis and root development did not show complete signals such as found in normal telomeres. These findings indicate that a certain time or stage is required for synthesis of the telomeric repetitive sequences with a complete length. Nevertheless, because the broken ends without complete telomere sequences were also healed, restoration of the normal complement of telomere sequences is not necessary for healing of broken ends.     

Metropolis Monte Carlo calculations of DNA structure using internal coordinates and NMR distance restraints: An alternative method for generating a high-resolution solution structure

Summary A new method, a restrained Monte Carlo (rMC) calculation, is demonstrated for generating high-resolution structures of DNA oligonucleotides in solution from interproton distance restraints and bounds derived from complete relaxation matrix analysis of two-dimensional nuclear Overhauser effect (NOE) spectral peak intensities. As in the case of restrained molecular dynamics (rMD) refinement of structures, the experimental distance restraints and bounds are incorporated as a pseudo-energy term (or penalty function) into the mathematical expression for the molecular energy. However, the use of generalized helical parameters, rather than Cartesian coordinates, to define DNA conformation increases efficiency by decreasing by an order of magnitude the number of parameters needed to describe a conformation and by simplifying the potential energy profile. The Metropolis Monte Carlo method is employed to simulate an annealing process. The rMC method was applied to experimental 2D NOE data from the octamer duplex d(GTA-TAATG)·d(CATTATAC). Using starting structures from different locations in conformational space (e.g. A-DNA and B-DNA), the rMC calculations readily converged, with a root-mean-square deviation (RMSD) of <0.3 Å between structures generated using different protocols and starting structures. Theoretical 2D NOE peak intensities were calculated for the rMC-generated structures using the complete relaxation matrix program CORMA, enabling a comparison with experimental intensities via residual indices. Simulation of the vicinal proton coupling constants was carried out for the structures generated, enabling a comparison with the experimental deoxyribose ring coupling constants, which were not utilized in the structure determination in the case of the rMC simulations. Agreement with experimental 2D NOE and scalar coupling data was good in all cases. The rMC structures are quite similar to that refined by a traditional restrained MD approach (RMSD<0.5 Å) despite the different force fields used and despite the fact that MD refinement was conducted with additional restraints imposed on the endocyclic torsion angles of deoxyriboses. The computational time required for the rMC and rMD calculations is about the same. A comparison of structural parameters is made and some limitations of both methods are discussed with regard to the average nature of the experimental restraints used in the refinement.Abbreviations MC Monte Carlo - rMC restrained Monte Carlo - MD molecular dynamics - rMD restrained molecular dynamics - DG distance geometry - EM energy minimization - 2D NOE two-dimensional nuclear Overhauser effect - DQF-COSY double-quantum-filtered correlation spectroscopy - RMSD root-mean-square deviation To whom correspondence should be addressed.     

Flower senescence in daylily (Hemerocallis)

This paper reviews recent developments in the use of molecular probes for analyzing the genetic makeup of somatic hybrids. Successful application of somatic hybridization to the interspecific transfer of traits encoded in the nucleus is still having limited success. A major difficulty is hybrid infertility, particularly in hybrids between sexually incompatible species. The formation of asymmetric hybrids is being explored as an approach for improving hybrid fertility. Evaluation of the degree of chromosome elimination and chromosome stability and instability in asymmetric hybrids is difficult when the traditional approaches of chromosome counting and isozyme analysis are used. Two new approaches are resolving this difficulty. The use of species-specific repetitive DNA probes in dot blotting and in situ hybridization to chromosomes is providing quantitative data on chromosome elimination and allows detection of translocations. Use of restriction fragment length polymorphism (RFLP) probes for analysis of hybrids between genetically mapped species makes it possible to account for the presence or absence of individual chromosomes and chromosomes arms. Wider use of such molecular probes should greatly improve our understanding of the genetics of both symmetric and asymmetric somatic hybrids and may lead to new strategies for the effective interspecific transfer of nucleus-encoded traits by protoplast fusion.