The 18S rRNA from Odontophrynus americanus 2n and 4n (Amphibia, Anura) reveals unusual extra sequences in the variable region V2.

The nucleotide sequence of the rDNA 18S region isolated from diploid and tetraploid species of the amphibian Odontophrynus americanus was determined and used to predict the secondary structure of the corresponding 18S rRNA molecules. Comparison of the primary and secondary structures for the 2n and 4n species confirmed that these species are very closely related. Only three nucleotide substitutions were observed, accounting for 99% identity between the 18S sequences, whereas several changes were detected by comparison with the Xenopus laevis 18S sequence (96% identity). Most changes were located in highly variable regions of the molecule. A noticeable feature of the Odontophrynus 18S rRNA was the presence of unusual extra sequences in the V2 region, between helices 9 and 11. These extra sequences do not fit the model for secondary structure predicted for vertebrate 18S rRNA.     

The nucleotide sequence and predicted secondary structure of small subunit (18S) ribosomal RNA from Spirometra erinaceieuropaei

The nucleotide (nt) sequence of a small subunit (18S) ribosomal RNA gene from the plerocercoid of Spirometra erinaceieuropaei (SEP) was determined. The gene with 2182 bp in length is larger than that of most eukaryotes. Extra nt sequences occur in regions known to be variable (V4 and V7). The predicted secondary structure of the nt positions 679–933 (V4) revealed different helices from that of other eukaryotes. The region between nt positions 1540 and 1749 (V7) was different from that of other eukaryotes, but the secondary structure prediction by computer analysis demonstrated that this part of 18S rRNA sequence from S. erinaceieuropaei may form a single extended helix. Nt that were aligned with those of nine other parasites were used to estimate phylogenetic relationships. The data presented here clearly indicate that S. erinaceieuropaei is closely related to Echinococcus granulosus.     

Evolution of Hypervariable Regions, V4 and V7, of Insect 18S rRNA and Their Phylogenetic Implications

We compared primary and secondary structures of V4 (helices E23-2 to E23-5) and V7 (helix 43) regions of 18S rRNAs in insects and the other three major arthropod groups (crustaceans, myriapods, and chelicerates) known so far. We found that the lengths of primary sequences and the shapes of secondary structures of these two hypervariable regions of insect 18S rRNA even at infraclass levels are phylogenetically informative and reflect major steps in insect evolution. The long sequence insertion and bifurcated shape of helices E23-2 to E23-5 in the V4 region are unique synapomorphic characters for winged insects (Pterygota). The long sequence insertion and expanded stem length of helix 43 in the V7 region are synapomorphic characters for holometabolous insects which conduct complete metamorphosis. The strongly conserved secondary structures suggest the possibility that these hypervariable regions may be related with certain important cellular functions unknown thus far. The comparison with insect fossil records revealed that the pterygote synapomorphy (V4) and the holometabolous synapomorphy (V7) were established prior to the acquisition of insect wings (flight system) and prior to the development of complete metamorphosis, respectively. These synapomorphies have been also relatively stable over at least 300 Myr and 280 Myr, respectively as well. It implies that the expansion events of the V4 and V7 regions have not occurred simultaneously but independently at different periods during the insect evolution. Then this suggests that V4 and V7 regions are not functionally correlated as recently suggested by Crease and Coulbourn.     

Putative secondary structures of unusually long strepsipteran SSU rRNAs and its phylogenetic implications.

We constructed the putative secondary structures of the small subunit rRNAs (SSU rRNA) from three strepsipteran insects. The primary sequences of the strepsipteran SSU rRNAs are unusually long due to unique and long insertions. In spite of these insertions, the basic shapes of their secondary structures are well maintained as shown in those of other eukaryotes, because these insertions appear mainly in the variable regions. The secondary structures for the V1, V3, V5, V8, and V9 regions are well conserved, even though the primary structures of V1, V5, and V8 regions are quite variable. However, the predicted secondary structures for the V2, V4, and V7 regions are quite different from those of other insects. In the V4 and V7 regions, helices specific to the Strepsiptera exist. These helices have not been reported in other organisms so far. Similarly, four eukaryotic specific helices (E8-1, E10-2, E23-4 and E45-1) not reported in insects exist in the V2, V4, and V8 regions. These helices are formed by the inserted sequences. The secondary structures of the expanded segments of the strepsipteran SSU rRNA were applied to infer the phylogenetic position of Strepsiptera, one of the most enigmatic problems in insect phylogeny. Only the secondary structure of the V7 region showed the weak Strepsiptera/Diptera sister-group relationship.     

A secondary structural model of the 28S rRNA expansion segments D2 and D3 for Chalcidoid wasps (Hymenoptera: Chalcidoidea)

We analyze the secondary structure of two expansion segments (D2, D3) of the 28S ribosomal (rRNA)-encoding gene region from 527 chalcidoid wasp taxa (Hymenoptera: Chalcidoidea) representing 18 of the 19 extant families. The sequences are compared in a multiple sequence alignment, with secondary structure inferred primarily from the evidence of compensatory base changes in conserved helices of the rRNA molecules. This covariation analysis yielded 36 helices that are composed of base pairs exhibiting positional covariation. Several additional regions are also involved in hydrogen bonding, and they form highly variable base-pairing patterns across the alignment. These are identified as regions of expansion and contraction or regions of slipped-strand compensation. Additionally, 31 single-stranded locales are characterized as regions of ambiguous alignment based on the difficulty in assigning positional homology in the presence of multiple adjacent indels. Based on comparative analysis of these sequences, the largest genetic study on any hymenopteran group to date, we report an annotated secondary structural model for the D2, D3 expansion segments that will prove useful in assigning positional nucleotide homology for phylogeny reconstruction in these and closely related apocritan taxa.     

Comparative study of the validity of three regions of the 18S‐rRNA gene for massively parallel sequencing‐based monitoring of the planktonic eukaryote community

The nuclear 18S‐rRNA gene has been used as a metabarcoding marker in massively parallel sequencing (MPS)‐based environmental surveys for plankton biodiversity research. However, different hypervariable regions have been used in different studies, and their utility has been debated among researchers. In this study, detailed investigations into 18S‐rRNA were carried out; we investigated the effective number of sequences deposited in international nucleotide sequence databases (INSDs), the amplification bias, and the amplicon sequence variability among the three variable regions, V1–3, V4–5 and V7–9, using in silico polymerase chain reaction (PCR) amplification based on INSDs. We also examined the primer universality and the taxonomic identification power, using MPS‐based environmental surveys in the Sea of Okhotsk, to determine which region is more useful for MPS‐based monitoring. The primer universality was not significantly different among the three regions, but the number of sequences deposited in INSDs was markedly larger for the V4–5 region than for the other two regions. The sequence variability was significantly different, with the highest variability in the V1–3 region, followed by the V7–9 region, and the lowest variability in the V4–5 region. The results of the MPS‐based environmental surveys showed significantly higher identification power in the V1–3 and V7–9 regions than in the V4–5 region, but no significant difference was detected between the V1–3 and V7–9 regions. We therefore conclude that the V1–3 region will be the most suitable for future MPS‐based monitoring of natural eukaryote communities, as the number of sequences deposited in INSDs increases.     

Characteristics of murine monoclonal anti-CD4. Epitope recognition, idiotype expression, and variable region gene sequence.

We have characterized a series of mouse monoclonal anti-CD4 and describe both their CD4 epitope recognition and Id expression. We also determined the V region gene sequences of these antibodies in an attempt to correlate epitope recognition and Id expression with V region sequence. All of these preparations recognize epitopes that cluster around the HIV gp120 binding site on the human CD4 molecule. However, we observed differences in epitope recognition among the anti-CD4 preparations, based on either competitive inhibition assays or functional assays, such as syncytium inhibition. Analysis of Id specificities using a polyclonal anti-Id generated against anti-Leu 3a indicated that five of the seven monoclonal anti-CD4 expressed a shared Id. Based on V region gene sequences, the V region kappa-chain (V[kappa]) from each of the seven antibodies was encoded by the V[kappa]21 gene family and expressed the J[kappa]4 gene segment. Those preparations that expressed the shared Id with anti-Leu 3a have virtually identical V[kappa] sequences, with a high degree of homology in the CDR. The VH region gene sequences of six of the seven antibodies also shared overall homology and appeared to be encoded by the J558 VH gene family. The seventh anti-CD4 VH region is encoded for by the VHGAM gene family. The majority of these antibodies used JH3 gene segment, although the JH2 and JH4 gene segments were also represented. In addition, several of these antibodies share a common sequence organization within their V-D-J joining regions that appears to involve N and P sequences to generate unique D segments. Together, these data suggest that differences in epitope recognition among the monoclonal anti-CD4 may reflect sequence variability primarily within the CDR3 region of both V[kappa] and VH. The basis for the detection of a shared Id most likely reflects the high degree of homology within the V[kappa] region sequences. In addition, these data, which are based on a limited analysis, suggest the possible restricted use of V region germ-line gene families in the secondary antibody response of BALB/c mice to specific epitopes on the human CD4 molecule.     

Identification and assignment of base pairs in three helical stems of Torulopsis utilis ribosomal 5S RNA and its RNase T1 and RNase T2 cleaved fragments via 500-MHz proton homonuclear overhauser enhancements

Imino proton resonances in the downfield region (10-14 ppm) of the 500-MHz 1H NMR spectrum of Torulopsis utilis 5S RNA are identified (A X U, G X C, or G X U) and assigned to base pairs in helices I, IV, and V via analysis of homonuclear Overhauser enhancements (NOE) from intact T. utilis 5S RNA, its RNase T1 and RNase T2 digested fragments, and a second yeast (Saccharomyces cerevisiae) 5S RNA whose nucleotide sequence differs at only six residues from that of T. utilis 5S RNA. The near-identical chemical shifts and NOE behavior of most of the common peaks from these four RNAs strongly suggest that helices I, IV, and V retain the same conformation after RNase digestion and that both T. utilis and S. cerevisiae 5S RNAs share a common secondary and tertiary structure. Of the four G X U base pairs identified in the intact 5S RNA, two are assigned to the terminal stem (helix I) and the other two to helices IV and V. Seven of the nine base pairs of the terminal stem have been assigned. Our experimental demonstration of a G X U base pair in helix V supports the 5S RNA secondary structural model of Luehrsen and Fox [Luehrsen, K. R., & Fox, G.E. (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 2150-2154]. Finally, the base-pair proton peak assigned to the terminal G X U in helix V of the RNase T2 cleaved fragment is shifted downfield from that in the intact 5S RNA, suggesting that helices I and V may be coaxial in intact T. utilis 5S RNA.     

18S rRNA secondary structure and phylogenetic position of Peloridiidae (Insecta, hemiptera)

A secondary structure model for 18S rRNA of peloridiids, relict insects with a present-day circumantarctic distribution, is constructed using comparative sequence analysis, thermodynamic folding, a consensus method using 18S rRNA models of other taxa, and support of helices based on compensatory substitutions. Results show that probable in vivo configuration of 18S rRNA is not predictable using current free-energy models to fold the entire molecule concurrently. This suggests that refinements in free-energy minimization algorithms are needed. Molecular phylogenetic datasets were created using 18S rRNA nucleotide alignments produced by CLUSTAL and rigorous interpretation of homologous position based on certain secondary substructures. Phylogenetic analysis of a hemipteran data matrix of 18S rDNA sequences placed peloridiids sister to Heteroptera. Resolution of affiliations between the three main euhemipteran lineages was unresolved. The peloridiid 18S RNA model presented here provides the most accurate template to date for aligning homologous nucleotides of hemipteran taxa. Using folded 18S rRNA to infer homology of character as morpho-molecular structures or nucleotides and scoring particular sites or substructures is discussed.     

A proposal for the secondary structure of a variable area of eukaryotic small ribosomal subunit RNA involving the existence of a pseudoknot.

Eukaryotic small ribosomal subunit RNAs contain an area of variable structure, V4, which comprises about 250 nucleotides in most species, whereas the corresponding area in bacterial small ribosomal subunit RNAs consists of about 64 nucleotides folded into a single hairpin. There is no consensus on the secondary structure of area V4 in eukaryotes, about 10 different models having been proposed. The prediction of a model on a comparative basis poses special problems because, due to the variability of the area in length as well as sequence, a dependable alignment is very difficult to achieve. A new model was derived by systematic examination of all combinations of helices that have been hitherto proposed, plus some new ones. The following properties of the helices were examined: transposability to all presently known sequences, presence of compensating substitutions, and thermodynamic stability. A model was selected by ranking all possible combinations of transposable helices according to the number of compensating substitutions scored. The optimal model comprises a pseudoknot and four hairpin structures. Certain species contain additional hairpins inserted between these structural elements, while in others the structure is partially or entirely deleted.     

A computer method for finding common base paired helices in aligned sequences: application to the analysis of random sequences.

We describe a new computer program that identifies conserved secondary structures in aligned nucleotide sequences of related single-stranded RNAs. The program employs a series of hash tables to identify and sort common base paired helices that are located in identical positions in more than one sequence. The program gives information on the total number of base paired helices that are conserved between related sequences and provides detailed information about common helices that have a minimum of one or more compensating base changes. The program is useful in the analysis of large biological sequences. We have used it to examine the number and type of complementary segments (potential base paired helices) that can be found in common among related random sequences similar in base composition to 16S rRNA from Escherichia coli. Two types of random sequences were analyzed. One set consisted of sequences that were independent but they had the same mononucleotide composition as the 16S rRNA. The second set contained sequences that were 80% similar to one another. Different results were obtained in the analysis of these two types of random sequences. When 5 sequences that were 80% similar to one another were analyzed, significant numbers of potential helices with two or more independent base changes were observed. When 5 independent sequences were analyzed, no potential helices were found in common. The results of the analyses with random sequences were compared with the number and type of helices found in the phylogenetic model of the secondary structure of 16S ribosomal RNA. Many more helices are conserved among the ribosomal sequences than are found in common among similar random sequences. In addition, conserved helices in the 16S rRNAs are, on the average, longer than the complementary segments that are found in comparable random sequences. The significance of these results and their application in the analysis of long non-ribosomal nucleotide sequences is discussed.     

Large expansion segments in 18S rDNA support a new sponge clade (Class Demospongiae, Order Haplosclerida)

Newly emerging molecular phylogenetic hypotheses involving the sponge Order Haplosclerida (Class Demospongiae) are far removed from traditional views on their classification using morphology. In the new grouping of marine haplosclerid taxa by molecular data all members of one highly supported clade were found to have three large indels in the 18S rRNA gene. These indels were not found in this gene in other marine haplosclerids or in any other demosponges analysed. These indels were found in the variable V4 and V7 region of the gene, had high GC contents and formed stable double stranded helices in the 18S rRNA secondary structure. These indels are very important synapomorphies, provide high support for an alternative taxonomic scheme and could help resolve the phylogeny of this order in conjunction with other phylogenetically informative characters.     

A unique secondary folding pattern for 5S RNA corresponds to the lowest energy homologous secondary structure in 17 different prokaryotes.

A general secondary structure is proposed for the 5S RNA of prokaryotic ribosomes, based on helical energy filtering calculations. We have considered all secondary structures that are common to 17 different prokaryotic 5S RNAs and for each 5S sequence calculated the (global) minimum energy secondary structure (300,000 common structures are possible for each sequence). The 17 different minimum energy secondary structures all correspond, with minor differences, to a single, secondary structure model. This is strong evidence that this general 5S folding pattern corresponds to the secondary structure of the functional 5S rRNA. The general 5S secondary structure is forked and in analogy with the cloverleaf of tRNA is named the "wishbone" model. It constant 8 double helical regions; one in the stem, four in the first, or constant arm, and three in the second arm. Four of these double helical regions are present in a model earlier proposed (1) and four additional regions not proposed by them are presented here. In the minimum energy general structure, the four helices in the constant arm are exactly 15 nucleotide pairs long. These helices are stacked in the sequences from gram-positive bacteria and probably stacked in gram-negative sequences as well. In sequences from gram-positive bacteria the length of the constant arm is maintained at 15 stacked pairs by an unusual minimum energy interaction involving a C26-G57 base pair intercalated between two adjacent helical regions.     

A hierarchy of RNA subdomains in assembly of the central domain of the 30 S ribosomal subunit

Beginning with the framework that has been developed for the assembly of the 30 S ribosomal subunit, we have identified a series of RNAs that are minimal binding sites for proteins S15, S6, S18, and S11 in the central domain from Thermus thermophilus. The minimal binding RNA for proteins S15, S6, and S18 consists of helix 22 and three-way junctions at both ends composed of portions of helices 20, 21, and 23. Addition of the remaining portion of helix 23 to this construct results in the minimal site for S11. Surprisingly, almost half of the central domain (helices 24, 25, and 26) is dispensable for binding the central domain proteins. Thus, at least two classes of RNA elements can be identified in ribosomal RNA. A protein-binding core element (such as helices 20, 21, 22, and 23) is required for the association of ribosomal proteins, whereas secondary binding elements (such as helices 24, 25, and 26) associate only with the preformed core RNP complex. Apparently, there may be a hierarchy of ribosomal RNA elements similar to the hierarchy of primary, secondary, and tertiary binding ribosomal proteins.     

The secondary structure of the unusually long 18S ribosomal RNA of the myxozoan Sphaerospora truttae and structural evolutionary trends in the Myxozoa

The nearly complete 18S rRNA sequence of the myxozoan parasite Sphaerospora truttae shows an extraordinary length (2,552bp) in comparison with other myxozoans and with metazoans in general (average 1,800-1,900bp). The sequence shows nucleotide insertions in most variable regions of the 18S rRNA (V2, V4, V5 and V7), with especially large expansion segments in V4 and V7. In the myxozoans, nucleotide insertions and specific secondary structures in these regions of the gene were found to be strongly related to large scale phylogenetic clustering and thus with the invertebrate host type. Whereas expansion segments were generally found to be absent in the malacasporeans and the clade of primary marine myxozoan species, they occur in all taxa of the clade containing freshwater species, where they showed a consistent secondary structure throughout. The longest expansion segments occur in S. truttae, Sphaerospora elegans and Leptotheca ranae, which represent a clade that has emerged after the malacosporeans and before the radiation of all other myxozoan genera. These three species demonstrate structural links to the malacosporeans as well as other unique features. A smaller number of nucleotide insertions in different subhelices and specific secondary structures appear to have evolved independently in two marine genera, i.e. Ceratomyxa and Parvicapsula. The secondary structural elements of V4 and V7 of the myxozoan 18S rRNAs were found to be highly informative and revealed evolutionary trends of various regions of the gene hitherto unknown, since previous analyses have been based on primary sequence data excluding these regions. Furthermore, the unique features of the V4 region in S. truttae allowed for the design of a highly specific PCR assay for this species.     

Nucleotide sequence of a mosquito 18S ribosomal RNA gene.

We have sequenced an 18S ribosomal RNA gene from the mosquito, Aedes albopictus. Computer alignment of the 1950 nucleotide coding region (56% A + T) with 18S rRNA sequences from two insect and three vertebrate species revealed greater sequence divergence among the insects than among the vertebrates. Sequence alignments showed that variable region V4, which has been considered to be the most poorly conserved domain in the 18S rRNA gene, was better conserved among insects and vertebrates than was the V6 domain.     

Gene conversion: a mechanism for generation of heterogeneity in the tprK gene of Treponema pallidum during infection

The tprK gene sequence of Treponema pallidum subspecies pallidum (T. pallidum) is heterogeneous within and among isolates. Heterogeneity in the tprK open reading frame is localized in seven discrete variable (V) regions, and variability results from apparent base changes, insertions or deletions. The TprK V regions are the focus of anti-TprK antibodies arising during infection. To test our hypothesis that V region sequences change during infection and passage, we developed a clonal isolate from the Chicago strain of T. pallidum and confirmed V region diversification during passage of this isolate. We describe the sequence anatomy of the seven V regions of tprK and the identification of putative donor sites for new V region sequences, and we propose a model for generation of new V regions by segmental gene conversion. These findings suggest that antigenic variation of TprK occurs in T. pallidum and may be important in immune evasion and persistence.     

The origin and evolution of variable-region helices in V4 and V7 of the small-subunit ribosomal RNA of branchiopod crustaceans

We sequenced the V4 and V7 regions of the small-subunit ribosomal RNA (SSU rRNA) from 38 species of branchiopod crustaceans (e.g., Artemia, Daphnia, Triops) representing all eight extant orders. Ancestral large-bodied taxa in the orders Anostraca, Notostraca, Laevicaudata, and Spinicaudata (limnadiids and cyzicids) possess the typical secondary structure in these regions, whereas the spinicaudatan Cyclestheria and all of the cladocerans (Anomopoda, Ctenopoda, Onychopoda, and Haplopoda) possess three unique helices. Although the lengths and primary sequences of the distal ends of these helices are extremely variable, their locations, secondary structures, and primary sequences at the proximal end are conserved, indicating that they are homologous. This evidence supports the classical view that Cladocera is a monophyletic group and that the cyclestheriids are transitional between spinicaudatans and cladocerans. The single origin and persistence since the Permian of the unique cladoceran helices suggests that births and deaths of variable region helices have been rare. The broad range of sequence divergences observed among the cladoceran helices permitted us to make inferences about their evolution. Although their proximal ends are very GC-biased, there is a significant negative correlation between length and GC content due to an increasing proportion of U at their distal ends. Slippage-like processes occurring at unpaired nucleotides or bulges, which are very U-biased, are associated with both helix origin and runaway length expansion. The overall GC contents and lengths of V4 and V7 are highly correlated. More surprisingly, the lengths of these SSU rRNA variable regions are also highly correlated with the length of the large-subunit rRNA expansion segment, D2, indicating that mechanisms affecting length variation do so both across single genes and across genes in the rRNA gene family.     

Structures of the transmembrane helices of the G-protein coupled receptor, rhodopsin.

An hypothesis is tested that individual peptides corresponding to the transmembrane helices of the membrane protein, rhodopsin, would form helices in solution similar to those in the native protein. Peptides containing the sequences of helices 1, 4 and 5 of rhodopsin were synthesized. Two peptides, with overlapping sequences at their termini, were synthesized to cover each of the helices. The peptides from helix 1 and helix 4 were helical throughout most of their length. The N- and C-termini of all the peptides were disordered and proline caused opening of the helical structure in both helix 1 and helix 4. The peptides from helix 5 were helical in the middle segment of each peptide, with larger disordered regions in the N- and C-termini than for helices 1 and 4. These observations show that there is a strong helical propensity in the amino acid sequences corresponding to the transmembrane domain of this G-protein coupled receptor. In the case of the peptides from helix 4, it was possible to superimpose the structures of the overlapping sequences to produce a construct covering the whole of the sequence of helix 4 of rhodopsin. As similar superposition for the peptides from helix 1 also produced a construct, but somewhat less successfully because of the disordering in the region of sequence overlap. This latter problem was more severe for helix 5 and therefore a single peptide was synthesized for the entire sequence of this helix, and its structure determined. It proved to be helical throughout. Comparison of all these structures with the recent crystal structure of rhodopsin revealed that the peptide structures mimicked the structures seen in the whole protein. Thus similar studies of peptides may provide useful information on the secondary structure of other transmembrane proteins built around helical bundles.     

Amino acid sequence analysis of the annexin super-gene family of proteins.

The annexins are a widespread family of calcium-dependent membrane-binding proteins. No common function has been identified for the family and, until recently, no crystallographic data existed for an annexin. In this paper we draw together 22 available annexin sequences consisting of 88 similar repeat units, and apply the techniques of multiple sequence alignment, pattern matching, secondary structure prediction and conservation analysis to the characterisation of the molecules. The analysis clearly shows that the repeats cluster into four distinct families and that greatest variation occurs within the repeat 3 units. Multiple alignment of the 88 repeats shows amino acids with conserved physicochemical properties at 22 positions, with only Gly at position 23 being absolutely conserved in all repeats. Secondary structure prediction techniques identify five conserved helices in each repeat unit and patterns of conserved hydrophobic amino acids are consistent with one face of a helix packing against the protein core in predicted helices a, c, d, e. Helix b is generally hydrophobic in all repeats, but contains a striking pattern of repeat-specific residue conservation at position 31, with Arg in repeats 4 and Glu in repeats 2, but unconserved amino acids in repeats 1 and 3. This suggests repeats 2 and 4 may interact via a buried saltbridge. The loop between predicted helices a and b of repeat 3 shows features distinct from the equivalent loop in repeats 1, 2 and 4, suggesting an important structural and/or functional role for this region. No compelling evidence emerges from this study for uteroglobin and the annexins sharing similar tertiary structures, or for uteroglobin representing a derivative of a primordial one-repeat structure that underwent duplication to give the present day annexins. The analyses performed in this paper are re-evaluated in the Appendix, in the light of the recently published X-ray structure for human annexin V. The structure confirms most of the predictions and shows the power of techniques for the determination of tertiary structural information from the amino acid sequences of an aligned protein family.