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1.
The members of the PKA regulatory subunit family (PKA-R family) were analyzed by multiple sequence alignment and clustering
based on phylogenetic tree construction. According to the phylogenetic trees generated from multiple sequence alignment of
the complete sequences, the PKA-R family was divided into four subfamilies (types I to IV). Members of each subfamily were
exclusively from animals (types I and II), fungi (type III), and alveolates (type IV). Application of the same methodology
to the cAMP-binding domains, and subsequently to the region delimited by β-strands 6 and 7 of the crystal structures of bovine
RIα and rat RIIβ (the phosphate-binding cassette; PBC), proved that this highly conserved region was enough to classify unequivocally
the members of the PKA-R family. A single signature sequence, F–G–E–[LIV]–A–L–[LIMV]–x(3)–[PV]–R–[ANQV]–A, corresponding to
the PBC was identified which is characteristic of the PKA-R family and is sufficient to distinguish it from other members
of the cyclic nucleotide-binding protein superfamily. Specific determinants for the A and B domains of each R-subunit type
were also identified. Conserved residues defining the signature motif are important for interaction with cAMP or for positioning
the residues that directly interact with cAMP. Conversely, residues that define subfamilies or domain types are not conserved
and are mostly located on the loop that connects α-helix B′ and β strand 7.
Received: 2 November 2000/Accepted: 14 June 2001 相似文献
2.
Benoit Cousineau Fabrice Leclerc Robert Cedergren 《Journal of molecular evolution》1997,45(6):661-670
Sequence similarity has given rise to the proposal that IF-2, EF-G, and EF-Tu are related through a common ancestor. We evaluate
this proposition and whether the relationship can be extended to other factors of protein synthesis. Analysis of amino acid
sequence similarity gives statistical support for an evolutionary affiliation among IF-1, IF-2, IF-3, EF-Tu, EF-Ts, and EF-G
and suggests further that this association is a result of gene duplication/fusion events. In support of this mechanism, the
three-dimensional structures of IF-3, EF-Tu, and EF-G display a predictable domain structure and overall conformational similarity.
The model that we propose consists of three consecutives duplication/fusion events which would have taken place before the
divergence of the three superkingdoms: eubacteria, archaea, and eukaryotes. The root of this protein superfamily tree would
be an ancestor of the modern IF-1 gene sequence. The repeated fundamental motif of this protein superfamily is a small RNA
binding domain composed of two α-helices packed along side of an antiparallel β-sheet.
Received: 17 October 1996 / Accepted: 10 June 1997 相似文献
3.
Evolution of the Integrin α and β Protein Families 总被引:4,自引:0,他引:4
Hughes AL 《Journal of molecular evolution》2001,52(1):63-72
A phylogenetic analysis of vertebrate and invertebrate α integrins supported the hypothesis that two major families of vertebrate
α integrins originated prior to the divergence of deuterostomes and protostomes. These two families include, respectively,
the αPS1 and αPS2 integrins of Drosophila melanogaster, and each family has duplicated repeatedly in vertebrates but not in Drosophila. In contrast, a third family (including αPS3) has duplicated in Drosophila but is absent from vertebrates. Vertebrate αPS1 and αPS2 family members are found on human chromosomes 2, 12, and 17. Linkage
of these family members may have been conserved since prior to the origin of vertebrates, and the two genes duplicated simultaneously.
A phylogenetic analysis of β integrins did not clearly resolve whether vertebrate β integrin genes duplicated prior to the
origin of vertebrates, although it suggested that at least the gene encoding vertebrate β4 may have done so. In general, the
phylogeny of neither α nor β integrins showed a close correspondence with patterns of α–β heterodimer formation or other functional
characteristics. One major exception to this trend involved αL, αM, αX, and αD, a monophyletic group of immune cell-expressed
α integrins, which share a number of common functional characteristics and have evolved in coordinated fashion with their
β integrin partners.
Received: 22 June 2000 / Accepted: 11 September 2000 相似文献
4.
Bocchetta M Gribaldo S Sanangelantoni A Cammarano P 《Journal of molecular evolution》2000,50(4):366-380
The phylogenetic placement of the Aquifex and Thermotoga lineages has been inferred from (i) the concatenated ribosomal proteins S10, L3, L4, L23, L2, S19, L22, and S3 encoded in
the S10 operon (833 aa positions); (ii) the joint sequences of the elongation factors Tu(1α) and G(2) coded by the str operon tuf and fus genes (733 aa positions); and (iii) the joint RNA polymerase β- and β′-type subunits encoded in the rpoBC operon (1130 aa positions). Phylogenies of r-protein and EF sequences support with moderate (r-proteins) to high statistical confidence (EFs) the placement of the two hyperthermophiles at the base of the bacterial clade
in agreement with phylogenies of rRNA sequences. In the more robust EF-based phylogenies, the branching of Aquifex and Thermotoga below the successive bacterial lineages is given at bootstrap proportions of 82% (maximum likelihood; ML) and 85% (maximum
parsimony; MP), in contrast to the trees inferred from the separate EF-Tu(1α) and EF-G(2) data sets, which lack both resolution
and statistical robustness. In the EF analysis MP outperforms ML in discriminating (at the 0.05 level) trees having A. pyrophilus and T. maritima as the most basal lineages from competing alternatives that have (i) mesophiles, or the Thermus genus, as the deepest bacterial radiation and (ii) a monophyletic A. pyrophilus–T. maritima cluster situated at the base of the bacterial clade. RNAP-based phylogenies are equivocal with respect to the Aquifex and Thermotoga placements. The two hyperthermophiles fall basal to all other bacterial phyla when potential artifacts contributed by the
compositionally biased and fast-evolving Mycoplasma genitalium and Mycoplasma pneumoniae sequences are eschewed. However, the branching order of the phyla is tenuously supported in ML trees inferred by the exhaustive
search method and is unresolved in ML trees inferred by the quartet puzzling algorithm. A rooting of the RNA polymerase-subunit
tree at the mycoplasma level seen in both the MP trees and the ML trees reconstructed with suboptimal amino acid substitution
models is not supported by the EF-based phylogenies which robustly affiliate mycoplasmas with low-G+C gram-positives and,
most probably, reflects a ``long branch attraction' artifact.
Received: 22 September 1999 / Accepted: 11 January 2000 相似文献
5.
Piero Cammarano Roberta Creti Anna M. Sanangelantoni Peter Palm 《Journal of molecular evolution》1999,49(4):524-537
A global alignment of EF-G(2) sequences was corrected by reference to protein structure. The selection of characters eligible
for construction of phylogenetic trees was optimized by searching for regions arising from the artifactual matching of sequence
segments unique to different phylogenetic domains. The spurious matchings were identified by comparing all sections of the
global alignment with a comprehensive inventory of significant binary alignments obtained by BLAST probing of the DNA and
protein databases with representative EF-G(2) sequences. In three discrete alignment blocks (one in domain II and two in domain
IV), the alignment of the bacterial sequences with those of Archaea–Eucarya was not retrieved by database probing with EF-G(2)
sequences, and no EF-G homologue of the EF-2 sequence segments was detected by using partial EF-G(2) sequences as probes in
BLAST/FASTA searches. The two domain IV regions (one of which comprises the ADP-ribosylatable site of EF-2) are almost certainly
due to the artifactual alignment of insertion segments that are unique to Bacteria and to Archaea–Eucarya. Phylogenetic trees
have been constructed from the global alignment after deselecting positions encompassing the unretrieved, spuriously aligned
regions, as well as positions arising from misalignment of the G′ and G″ subdomain insertion segments flanking the ``fifth'
consensus motif of the G domain (?varsson, 1995). The results show inconsistencies between trees inferred by alternative methods
and alternative (DNA and protein) data sets with regard to Archaea being a monophyletic or paraphyletic grouping. Both maximum-likelihood
and maximum-parsimony methods do not allow discrimination (by log-likelihood difference and difference in number of inferred
substitutions) between the conflicting (monophyletic vs. paraphyletic Archaea) topologies. No specific EF-2 insertions (or
terminal accretions) supporting a crenarchaeal–eucaryal clade are detectable in the new EF-G(2) sequence alignment. 相似文献
6.
Hiroshi Wada Mari Kobayashi Riki Sato Nori Satoh Hitoshi Miyasaka Yoshihisa Shirayama 《Journal of molecular evolution》2002,54(1):118-128
To test the validity of intron–exon structure as a phylogenetic marker, the intron–exon structure of EF-1α genes was investigated
for starfish, acornworms, ascidians, larvaceans, and amphioxus and compared with that of vertebrates. Of the 11 distinct intron
insertion sites found within the coding regions of the deuterostome EF-1α genes, 7 are shared by several taxa, while the remainder
are unique to certain taxa. Examination of the shared introns of the deuterostome EF-1α gene revealed that independent intron
loss or intron insertion must have occurred in separate lineages of the deuterostome taxa. Maximum parsimony analysis of the
intron–exon data matrix recovered five parsimonious trees (consistency index = 0.867). From this result, we concluded that
the intron–exon structure of deuterostome EF-1α has evolved more dynamically than previously thought, rendering it unsuitable
as a phylogenetic marker. We also reconstructed an evolutionary history of intron insertion–deletion events on the deuterostome
phylogeny, based on several molecular phylogenetic studies. These analyses revealed that the deuterostome EF-1α gene has lost
individual introns more frequently than all introns simultaneously. 相似文献
7.
The available amino acid sequences of the α-amylase family (glycosyl hydrolase family 13) were searched to identify their
domain B, a distinct domain that protrudes from the regular catalytic (β/α)8-barrel between the strand β3 and the helix α3. The isolated domain B sequences were inspected visually and also analyzed
by Hydrophobic Cluster Analysis (HCA) to find common features. Sequence analyses and inspection of the few available three-dimensional
structures suggest that the secondary structure of domain B varies with the enzyme specificity. Domain B in these different
forms, however, may still have evolved from a common ancestor. The largest number of different specificities was found in
the group with structural similarity to domain B from Bacillus cereus oligo-1,6-glucosidase that contains an α-helix succeeded by a three-stranded antiparallel β-sheet. These enzymes are α-glucosidase,
cyclomaltodextrinase, dextran glucosidase, trehalose-6-phosphate hydrolase, neopullulanase, and a few α-amylases. Domain B
of this type was observed also in some mammalian proteins involved in the transport of amino acids. These proteins show remarkable
similarity with (β/α)8-barrel elements throughout the entire sequence of enzymes from the oligo-1,6-glucosidase group. The transport proteins, in
turn, resemble the animal 4F2 heavy-chain cell surface antigens, for which the sequences either lack domain B or contain only
parts thereof. The similarities are compiled to indicate a possible route of domain evolution in the α-amylase family.
Received: 4 December 1996 / Accepted: 13 March 1997 相似文献
8.
Mariana Mondragón-Palomino Daniel Piñero Anne Nicholson-Weller Juan P. Laclette 《Journal of molecular evolution》1999,49(2):282-289
The plasma complement system comprises several activation pathways that share a common terminal route involving the assembly
of the terminal complement complex (TCC), formed by C5b–C9. The order of emergence of the homologous components of TCC (C6,
C7, C8α, C8β, and C9) has been determined by phylogenetic analyses of their amino acid sequences. Using all the sequence data
available for C6–C9 proteins, as well as for perforins, the results suggested that these TCC components originated from a
single ancestral gene and that C6 and C7 were the earliest to emerge. Our evidence supports the notion that the ancestral
gene had a complex modular composition. A series of gene duplications in combination with a tendency to lose modules resulted
in successive complement proteins with decreasing modular complexity. C9 and perforin apparently are the result of different
selective conditions to acquire pore-forming function. Thus C9 and perforin are examples of evolutionary parallelism.
Received: 16 August 1998 / Accepted: 12 March 1999 相似文献
9.
To test sodium channel structural models, we defined the epitopes for nineteen independently cloned monoclonal antibodies
previously generated against purified, detergent-solubilized, adult rat skeletal muscle sodium channel protein using channel
proteolysis, synthetic peptides, and fusion proteins. All identified epitopes were continuous and unique to the skeletal muscle
subtype α-subunit. Of the nineteen independent clones, seventeen had epitopes located either in the origin of the amino-terminus
or in the interdomain 2–3 region while only two antibodies had epitopes located in the mid-portion of the interdomain 1–2
region. No immunogenic regions were identified on the α-subunit's extracellular regions, interdomain 3–4 segment, or carboxyl-terminus
or on channel β-subunits. While immune tolerance may explain the lack of immunogenicity of extracellular regions, the lack
of immunogenicity of most of the channel's cytoplasmic mass may be due to segment inaccessibility from organization of these
regions as globular domains, to insertion of parts of these regions into the membrane phase, or to interaction with other
protein elements. The definition of monoclonal antibody epitopes allows us to reinterpret previously reported monoclonal antibody
competition studies, providing independent support for our model of sodium channel cytoplasmic domain structure. In addition,
these data suggest additional testable hypotheses concerning the interactions of the sodium channel amino- and carboxyl-termini
with each other as well as with other protein elements.
Received: 4 March 1998/Revised: 15 May 1998 相似文献
10.
Quantitative analyses were carried out on a large number of proteins that contain the highly conserved basic helix–loop–helix
domain. Measures derived from information theory were used to examine the extent of conservation at amino acid sites within
the bHLH domain as well as the extent of mutual information among sites within the domain. Using the Boltzmann entropy measure,
we described the extent of amino acid conservation throughout the bHLH domain. We used position association (pa) statistics that reflect the joint probability of occurrence of events to estimate the ``mutual information content' among
distinct amino acid sites. Further, we used pa statistics to estimate the extent of association in amino acid composition at each site in the domain and between amino acid
composition and variables reflecting clade and group membership, loop length, and the presence of a leucine zipper. The pa values were also used to describe groups of amino acid sites called ``cliques' that were highly associated with each other.
Finally, a predictive motif was constructed that accurately identifies bHLH domain-containing proteins that belong to Groups
A and B.
Received: 15 December 1997 / Accepted: 1 October 1998 相似文献
11.
The amino acid sequences of 22 α-amylases from family 13 of glycosyl hydrolases were analyzed with the aim of revealing the
evolutionary relationships between the archaeal α-amylases and their eubacterial and eukaryotic counterparts. Two evolutionary
distance trees were constructed: (i) the first one based on the alignment of extracted best-conserved sequence regions (58
residues) comprising β2, β3, β4, β5, β7, and β8 strand segments of the catalytic (α/β)8-barrel and a short conserved stretch in domain B protruding out of the barrel in the β3 →α3 loop, and (ii) the second one
based on the alignment of the substantial continuous part of the (α/β)8-barrel involving the entire domain B (consensus length: 386 residues). With regard to archaeal α-amylases, both trees compared
brought, in fact, the same results; i.e., all family 13 α-amylases from domain Archaea were clustered with barley pI isozymes,
which represent all plant α-amylases. The enzymes from Bacillus licheniformis and Escherichia coli, representing liquefying and cytoplasmic α-amylases, respectively, seem to be the further closest relatives to archaeal α-amylases.
This evolutionary relatedness clearly reflects the discussed similarities in the amino acid sequences of these α-amylases,
especially in the best-conserved sequence regions. Since the results for α-amylases belonging to all three domains (Eucarya,
Eubacteria, Archaea) offered by both evolutionary trees are very similar, it is proposed that the investigated conserved sequence
regions may indeed constitute the ``sequence fingerprints' of a given α-amylase.
Received: 3 June 1998 / Accepted: 20 August 1998 相似文献
12.
Sequences from the tuf gene coding for the elongation factor EF-Tu were amplified and sequenced from the genomic DNA of Pirellula marina and Isosphaera pallida, two species of bacteria within the order Planctomycetales. A near-complete (1140-bp) sequence was obtained from Pi. marina and a partial (759-bp) sequence was obtained for I. pallida. Alignment of the deduced Pi. marina EF-Tu amino acid sequence against reference sequences demonstrated the presence of a unique 11-amino acid sequence motif
not present in any other division of the domain Bacteria. Pi. marina shared the highest percentage amino acid sequence identity with I. pallida but showed only a low percentage identity with other members of the domain Bacteria. This is consistent with the concept
of the planctomycetes as a unique division of the Bacteria. Neither primary sequence comparison of EF-Tu nor phylogenetic
analysis supports any close relationship between planctomycetes and the chlamydiae, which has previously been postulated on
the basis of 16S rRNA. Phylogenetic analysis of aligned EF-Tu amino acid sequences performed using distance, maximum-parsimony,
and maximum-likelihood approaches yielded contradictory results with respect to the position of planctomycetes relative to
other bacteria. It is hypothesized that long-branch attraction effects due to unequal evolutionary rates and mutational saturation
effects may account for some of the contradictions.
Received: 21 August 2000 / Accepted: 8 January 2001 相似文献
13.
Sakai H Imamura C Osada Y Saito R Washio T Tomita M 《Journal of molecular evolution》2001,52(2):164-170
In this study, we analyzed the correlation between codon usage bias and Shine–Dalgarno (SD) sequence conservation, using
complete genome sequences of nine prokaryotes. For codon usage bias, we adopted the codon adaptation index (CAI), which is
based on the codon usage preference of genes encoding ribosomal proteins, elongation factors, heat shock proteins, outer membrane
proteins, and RNA polymerase subunit proteins. To compute SD sequence conservation, we used SD motif sequences predicted by
Tompa and systematically aligned them with 5′UTR sequences. We found that there exists a clear correlation between the CAI
values and SD sequence conservation in the genomes of Escherichia coli, Bacillus subtilis, Haemophilus influenzae, Archaeoglobus fulgidus, Methanobacterium thermoautotrophicum, and Methanococcus jannaschii, and no relationship is found in M. genitalium, M. pneumoniae, and Synechocystis. That is, genes with higher CAI values tend to have more conserved SD sequences than do genes with lower CAI values in these
organisms. Some organisms, such as M. thermoautotrophicum, do not clearly show the correlation. The biological significance of these results is discussed in the context of the translation
initiation process and translation efficiency.
Received: 22 June 2000 / Accepted: 18 October 2000 相似文献
14.
Díez-Sampedro A Lostao MP Wright EM Hirayama BA 《The Journal of membrane biology》2000,176(2):111-117
Using cotransporters as drug delivery vehicles is a topic of continuing interest. We examined glucose derivatives containing
conjugated aromatic rings using two isoforms of the Na+/glucose cotransporter: human SGLT1 (hSGLT1) and pig SGLT3 (pSGLT3, SAAT1). Our studies indicate that there is similarity
between SGLT1 and SGLT3 in the overall architecture of the vestibule leading to the sugar-binding site but differences in
translocation pathway interactions. Indican was transported by hSGLT1 with higher affinity (K0.5 0.06 mm) and 2-naphthylglucose with lower affinity (K0.5 0.5 mm) than α-methyl-d-glucopyranoside (αMDG, 0.2 mm). Both were poorly transported (maximal velocities, I
max
, 14% and 8% of αMDG). Other compounds were inhibitors (K
i
s 1–13 mm). In pSGLT3, indican and 2-naphthylglucose were transported with higher affinity than αMDG (K0.5s 0.9, 0.2 and 2.5 mm and relative I
max
s of 80, 25 and 100%). Phenylglucose and arbutin were transported with higher I
max
s (130 and 120%) and comparable K0.5s (8 and 1 mm). Increased affinity of indican relative to αMDG suggests that nitrogen in the pyrrole ring is favorable in both transporters.
Higher affinity of 2-naphthylglucose for pSGLT3 than hSGLT1 suggests more extensive hydrophobic/aromatic interaction in pSGLT3
than in hSGLT1. Our results indicate that bulky hydrophobic glucosides can be transported by hSGLT1 and pSGLT3, and discrimination
between them is based on steric factors and requirements for H-bonding. This provides information for design of glycosides
with potential therapeutic value.
Received: 18 February 2000/Revised: 13 April 2000 相似文献
15.
The major facilitator superfamily (MFS) of transport proteins, which includes the lactose permease of Escherichia coli, contains a conserved motif G-X-X-X-D/E-R/K-X-G-R/K-R/K in the loops that connect transmembrane segments 2 and 3, and transmembrane
segments 8 and 9. In three previous studies (Jessen-Marshall, A.E., & Brooker, R.J. 1996. J. Biol. Chem.
271:1400–1404; Jessen-Marshall, A.E., Parker, N., & Brooker, R.J. 1997. J. Bacteriol.
179:2616–2622; and Pazdernik, N., Cain, S.M., & Brooker, R.J. 1997. J. Biol. Chem.
272:26110–26116), suppressor mutations at twenty different sites were identified which restore function to mutant permeases that
have deleterious mutations in the conserved loop 2/3 or loop 8/9 motif. In the current study, several of these second-site
suppressor mutations have been separated from the original mutation in the conserved motif. The loop 2/3 suppressors were
then coupled to a loop 8/9 mutation (P280L) and the loop 8/9 suppressors were coupled to a loop 2/3 mutation (i.e., G64S)
to determine if the suppressors could restore function only to a loop 2/3 mutation, a loop 8/9 mutation, or both.
The single parent mutations changing the first position in loop 2/3 (i.e., G64S) and loop 8/9 (i.e., P280L) had less than
4% lactose transport activity. Interestingly, most of the suppressors were very inhibitory when separated from the parent
mutation. Two suppressors, A50T and G370V, restored substantial transport activity when individually coupled to the mutation
in loop 2/3 and also when coupled to the corresponding mutation in loop 8/9. In other words, these suppressors could alleviate
a defect imposed by mutations in either half of the permease. From a kinetic analysis, these suppressors were shown to exert
their effects by increasing the V
max
values for lactose transport compared with the single G64S and P280L strains. These results are discussed within the context
of our model in which the two halves of the lactose permease interact at a rotationally symmetrical interface, and that lactose
transport is mediated by conformational changes at the interface.
Received: 18 November 1999/Revised: 11 April 2000 相似文献
16.
Recent finding that a prokaryote synthesizes lysine through the α-aminoadipate pathway demonstrates that the lysine synthesis
through the α-aminoadipate pathway is not typical of fungi. However, the fungal lysine biosynthesis is not completely the
same as the prokaryotic one. We point out that α-aminoadipate reductase is a key enzyme to the evolution of fungal lysine
synthesis. In addition, fungi have two different saccharopine dehydrogenases, which is also characteristic of fungi.
Received: 18 February 2000 / Accepted: 19 June 2000 相似文献
17.
Bouzat JL McNeil LK Robertson HM Solter LF Nixon JE Beever JE Gaskins HR Olsen G Subramaniam S Sogin ML Lewin HA 《Journal of molecular evolution》2000,51(6):532-543
We employed a phylogenomic approach to study the evolution of α subunits of the proteasome gene family from early diverging
eukaryotes. BLAST similarity searches of the Giardia lamblia genome identified all seven α proteasome genes characteristic of eukaryotes from the crown group. In addition, a PCR strategy
for the amplification of multiple α subunit sequences generated single α proteasome products for representatives of the Kinetoplastida
(Leishmania major), the Parabasalia (Trichomonas vaginalis), and the Microsporidia (Vairimorpha sp., Nosema sp., Endoreticulata sp., and Spraguea lophii). The kinetoplastid Trypanosoma cruzi and the eukaryote crown group Acanthamoeba castellanii yielded two distinct α proteasome genes each. The presence of seven distinct α proteasome genes in G. lamblia, one of the earliest-diverging eukaryotes, indicates that the α proteasome gene family evolved rapidly from a minimum of one
gene in Archaea to seven or more in Eukarya. Results from the phylogenomic analysis are consistent with the idea that the
Diplomonida (as represented by G. lamblia), the Kinetoplastida, the Parabasalia, and the Microsporidia diverged after the duplication events that originated the α
proteasome gene family. A model for the early origin and evolution of the proteasome gene family is presented.
Received: 14 February 2000 / Accepted: 14 August 2000 相似文献
18.
M.L. Tierney B. Birnir N.P. Pillai J.D. Clements S. M. Howitt G. B. Cox P.W. Gage 《The Journal of membrane biology》1996,154(1):11-21
The conserved leucine residues at the 9′ positions in the M2 segments of α1 (L264) and β1 (L259) subunits of the human GABAA receptor were replaced with threonine. Normal or mutant α1 subunits were co-expressed with normal or mutant β1 subunits in Sf9 cells using the baculovirus/Sf9 expression system. Cells in which one or both subunits were mutated had a
higher ``resting' chloride conductance than cells expressing wild-type α1β1 receptors. This chloride conductance was blocked by 10 mm penicillin, a recognized blocker of GABAA channels, but not by bicuculline (100 μm) or picrotoxin (100 μm) which normally inhibit the chloride current activated by GABA: nor was it potentiated by pentobarbitone (100 μm). In cells expressing wild-type β1 with mutated α1 subunits, an additional chloride current could be elicited by GABA but the rise time and decay were slower than for wild-type
α1β1 receptors. In cells expressing mutated β1 subunits with wild-type or mutated α1 subunits (αβ(L9′T) and α(L9′T)β(L9′T)), no response to GABA could be elicited: this was not due to an absence of GABAA receptors in the plasmalemma because the cells bound [3H]-muscimol. It was concluded that in GABAA channels containing the L9′T mutation in the β1 subunit, GABA-binding does not cause opening of channels, and that the L9′T mutation in either or both subunits gives an
open-channel state of the GABAA receptor in the absence of ligand.
Received: 17 April 1996/Revised: 5 July 1996 相似文献
19.
Charles Robin Robyn J. Russell Kerrie M. Medveczky John G. Oakeshott 《Journal of molecular evolution》1996,43(3):241-252
The α-esterase cluster of D. melanogaster contains 11 esterase genes dispersed over 60 kb. Embedded in the cluster are two unrelated open reading frames that have
sequence similarity with genes encoding ubiquitin-conjugating enzyme and tropomyosin. The esterase amino acid sequences show
37–66% identity with one another and all but one have all the motifs characteristic of functional members of the carboxyl/cholinesterase
multigene family. The exception has several frameshift mutations and appears to be a pseudogene. Patterns of amino acid differences
among cluster members in relation to generic models of carboxyl/cholinesterase protein structure are broadly similar to those
among other carboxyl/cholinesterases sequenced to date. However the α-esterases differ from most other members of the family
in: their lack of a signal peptide; the lack of conservation in cysteines involved in disulfide bridges; and in four indels,
two of which occur in or adjacent to regions that align with proposed substrate-binding sites of other carboxyl/cholinesterases.
Phylogenetic analyses clearly identify three simple gene duplication events within the cluster. The most recent event involved
the pseudogene which is located in an intron of another esterase gene. However, relative rate tests suggest that the pseudogene
remained functional after the duplication event and has become inactive relatively recently. The distribution of indels also
suggests a deeper node in the gene phylogeny that separates six genes at the two ends of the cluster from a block of five
in the middle.
Received: 18 January 1996 / Accepted: 12 March 1996 相似文献
20.
Isoform-Specific Function and Distribution of Na/K Pumps in the Frog Lens Epithelium 总被引:3,自引:1,他引:2
Epithelial cells from the anterior and equatorial surfaces of the frog lens were isolated and used the same day for studies
of the Na/K ATPase. RNase protection assays showed that all cells express α1- and α2-isoforms of the Na/K pump but not the α3-isoform, however the α2-isoform dominates in anterior cells whereas the α1-isoform dominates in equatorial cells. The whole cell patch-clamp technique was used to record functional properties of the
Na/K pump current (I
P
), defined as the current specifically inhibited by dihydro-ouabain (DHO). DHO-I
P
blockade data indicate the α1-isoform has a dissociation constant of 100 μm DHO whereas for the α2-isoform it is 0.75 μm DHO. Both α1- and α2-isoforms are half maximally activated at an intracellular Na+-concentration of 9 mm. The α1-isoform is half maximally activated at an extracellular K+-concentration of 3.9 mm whereas for the α2-isoform, half maximal activation occurs at 0.4 mm. Lastly, transport by the α1-isoform is inhibited by a drop in extracellular pH, which does not affect transport by the α2-isoform. Under normal physiological conditions, I
P
in equatorial cells is approximately 0.23 μA/μF, and in anterior cells it is about 0.14 μA/μF. These current densities refer
to the area of cell membrane assuming a capacitance of around 1 μF/cm2. Because cell size and geometry are different at the equatorial vs. anterior surface of the intact lens, we estimate Na/K pump current density per area of lens surface to be around 10 μA/cm2 at the equator vs. 0.5 μA/cm2 at the anterior pole.
Received: 17 May 2000/Revised: 11 August 2000 相似文献