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Kinematics of vertical climbing in lorises and Cheirogaleus medius.

The type of climbing exhibited by apes and atelines is argued to have been important in the evolution of specialized locomotion, such as suspensory locomotion and bipedalism. However, little is known about the mechanics of climbing in primates. Previous work shows that Asian apes and atelines use larger joint excursions and longer strides than African apes and the Japanese macaque, respectively. This study expands knowledge of climbing mechanics by providing the first quantitative kinematic data for vertical climbing in four prosimian species: three lorisid species (Loris tardigradus, Nycticebus coucang, and Nycticebus pygmaeus) that share with apes and atelines morphological traits arguably related to climbing, and a more generalized quadruped, Cheirogaleus medius. Subjects were videotaped as they climbed up a wooden pole. Kinematic values, such as step length and limb excursions, were calculated and compared between species. The results of this study show that lorises, like Asian apes and spider monkeys, use relatively larger joint excursions and longer steps than does C. medius during climbing. These data lend further support to the idea that some primate species (e.g., lorises, atelines, and apes) are more specialized kinematically and morphologically for climbing than others. Pilot data suggest that such kinematic differences in climbing style across broad phylogenetic groups may relate to the energetics of climbing. Such data may be important for understanding the morphological and kinematic adaptations to climbing exhibited by some primates.

Adaptation, Physiological↗

Survival and local recurrence after anterior resection and abdominoperineal excision for rectal cancer.

The aim of this retrospective study is to compare the outcome of abdominoperineal excision (APE) and anterior resection (AR) for rectal cancer in 136 patients. Local recurrence rates and 5-year survival probabilities were estimated for the AR and APE group. Further comparisons were carried out between hand-sewn and stapled anastomoses after AR, and between patients after AR and APE for tumours 2 to 6 cm from the dentate line. Local recurrence after AR occurred in 14% and after APE in 10% of these cases. Five-year survival probabilities and local recurrence frequencies showed no statistically significant difference (P > 0.05). Local recurrence rates were 13.5% after hand-sewn anastomoses and 15% after the stapled procedure. No statistically significant difference was observed in the 5-year survival and recurrence rate (P > 0.05). Seventy-four of 136 patients had tumours located 2 to 6 cm from the dentata line. Local recurrence occurred in 21% after AR and 5% after APE, showing a statistically significant difference in frequency (P < 0.05). No significant difference was found in cumulative 5-year survival probabilities. APE for advanced low rectal cancer showed a significant reduction in local recurrences compared to AR.

Humans↗

Isolation of aryl-phosphate ester-degrading bacterium from leachate of a sea-based waste disposal site.

An aryl-phosphate ester (APE)-degrading bacterium was isolated from the leachate of a sea-based waste disposal site. The isolated APE-degrading bacterial strain YS-57 grew well in a medium containing glucose and NaCl, and degraded two types of APE:tricresyl phosphate and triphenyl phosphate. The optimal temperature, pH, and NaCl concentration for the growth of strain YS-57 were 30 degrees C, 7.0, and 1.0%, respectively. Strain YS-57 grew at an APE concentration of 25 mg/l without being inhibited. APEs were degraded by the supernatant of the medium in which strain YS-57 was cultured, suggesting that the APE-degrading enzyme was released into the extracellular space in the logarithmic growth phase. The 16S rDNA sequence of strain YS-57 showed 95.6% similarity to that of Roseobacter gallaeciensis and the morphological properties were also comparable. Consequently, strain YS-57 was closely related to the genus Roseobacter.

Journal Article↗

NMR studies of abasic sites in DNA duplexes: deoxyadenosine stacks into the helix opposite acyclic lesions.

Proton and phosphorus NMR studies are reported for two complementary nonanucleotide duplexes containing acyclic abasic sites. The first duplex, d(C-A-T-G-A-G-T-A-C).d(G-T-A-C-P-C-A-T-G), contains an acyclic propanyl moiety, P, located opposite a deoxyadenosine at the center of the helix (designated APP 9-mer duplex). The second duplex, d(C-A-T-G-A-G-T-A-C).d(G-T-A-C-E-C-A-T-G), contains a similarly located acyclic ethanyl moiety, E (designated APE 9-mer duplex). The ethanyl moiety is one carbon shorter than the natural carbon-phosphodiester backbone of a single nucleotide unit of DNA. The majority of the exchangeable and nonexchangeable base and sugar protons in both the APP 9-mer and APE 9-mer duplexes, including those at the abasic site, have been assigned by recording and analyzing two-dimensional phase-sensitive NOESY data sets in H2O and D2O solution between -5 and 5 degrees C. These spectroscopic observations establish that A5 inserts into the helix opposite the abasic site (P14 and E14) and stacks between the flanking G4.C15 and G6.C13 Watson-Crick base pairs in both the APP 9-mer and APE 9-mer duplexes. The helix is right-handed at and adjacent to the abasic site, and all glycosidic torsion angles are anti in both 9-mer duplexes. Proton NMR parameters for the APP 9-mer and APE 9-mer duplexes are similar to those reported previously for the APF 9-mer duplex (F = furan) in which a cyclic analogue of deoxyribose was embedded in an otherwise identical DNA sequence [Kalnik, M. W., Chang, C. N., Grollman, A. P., & Patel, D. J. (1988) Biochemistry 27, 924-931]. These proton NMR experiments demonstrate that the structures at abasic sites are very similar whether the five-membered ring is open or closed or whether the phosphodiester backbone is shortened by one carbon atom. Phosphorus spectra of the APP 9-mer and APE 9-mer duplexes (5 degrees C) indicate that the backbone conformation is similarly perturbed at three phosphodiester backbone torsion angles. These same torsion angles are also distorted in the APF 9-mer but assume a different conformation than those in the APP 9-mer and APE 9-mer duplexes.

Base Composition↗

Quantum simulation of a hydrated noradrenaline analog with the torsional path integral method.

An extended version of the torsional path integral Monte Carlo (TPIMC) method is presented and shown to be useful for studying the conformation of flexible molecules in solvated clusters. The new technique is applied to the hydrated clusters of the 2-amino-1-phenyl-ethanol (APE) molecule. APE + nH2O clusters with n = 0-4 are studied at 100 and 300 K using both classical and quantum simulations. Only at the lower temperature is the hydration number n found to impact the conformational distribution of the APE molecule. This is shown to be a result of the temperature-dependent balance between the internal energy and entropy contributions to the relative conformer free energies. Furthermore, at 100 K, large quantum effects are observed in the calculated conformer populations. A particularly large quantum shift of 30% of the total population is calculated for the APE + 2H2O cluster, which is explained in terms of the relative zero point energy of the lowest-energy hydrated structures for this cluster. Finally, qualitative agreement is found between the reported calculations and recent spectroscopy experiments on the hydrated clusters of APE, including an entropically driven preference for the formation of AG-type hydrated structures and the formation of a water "droplet" in the APE + 4H2O cluster.

Models, Molecular↗

Strategies used to combine seriated cups by chimpanzees (Pan troglodytes), bonobos (Pan paniscus), and capuchins (Cebus apella).

The authors investigated strategies used to combine seriated cups by apes (Pan troglodytes and P. paniscus) and monkeys (Cebus apella) using a protocol reported in P. M. Greenfield, K. Nelson, and E. Saltzman's (1972) study with children. It was hypothesized that apes would exhibit more hierarchical combinations of cups than monkeys, given apes' language capacity, and that apes would seriate the cups more efficiently than monkeys. As predicted, apes made many structures with the cups using a variety of strategies, and monkeys rarely combined the cups. After a training phase to orient monkeys to the task, the 2 genera did not differ in the strategies used to combine the cups or in efficiency in seriating the cups. Success in this task suggests that sensorimotor versions of hierarchically organized combinatorial activity are well within apes' and monkeys' abilities.

Animals↗

Photoaffinity labeling with 2(-)[2-(4-azido-3(-)[125I]- iodophenyl)ethylamino]adenosine and autoradiography with 2(-)[2-(4-amino-3(-)[125I]iodophenyl)ethylamino]adenosine of A2a adenosine receptors in rat brain.

The A2a adenosine receptor agonist 2(-)[2-(4-amino-3- iodophenyl)ethylamino]adenosine is a potent coronary vasodilator. The corresponding radioiodinated ligand, [125I]APE, discriminates between high- and low-affinity conformations of A2a adenosine receptors. In this study, [125I]APE was used for rapid (24-h) autoradiography in rat brain sections. The pattern of [125I]APE binding is consistent with that expected of an A2a-selective radioligand. It is highest in striatum, nucleus accumbens, and olfactory tubercle, with little binding to cortex and septal nuclei. Specific [125I]APE binding to these brain regions is abolished by 1 microM 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine (CGS-21680) but is little affected by 100 nM 8-cyclopentyl-1,3-dipropylxanthine. Conversion of [125I]APE to the corresponding arylazide results in [125I]AzPE. The rank-order potency of compounds to compete for [125I]AzPE binding in the dark is CGS-21680 > D-(R)-N6-phenylisopropyladenosine > N6- cyclopentyladenosine, indicating that it also is an A2a-selective ligand. Specific photoaffinity labeling by [125I]AzPE of a single polypeptide (42 kDa) corresponding to A2a adenosine receptors is reduced 55 +/- 4% by 100 microM guanosine 5'-O-(3-thiotriphosphate) and 91 +/- 1.3% by 100 nM CGS-21680. [125I]APE and [125I]AzPE are valuable new tools for characterizing A2a adenosine receptors and their coupling to GTP-binding proteins by autoradiography and photoaffinity labeling.

Adenosine↗

Natural antibodies in sera from healthy humans to antigens on surfaces of type C RNA viruses and cells from primates.

Sera from healthy humans contained naturally occurring antibody against group- or subgroup-specific antigen on the envelope of the following type C viruses isolated from primates: gibbon ape leukemia virus, simian (woolly monkey) sarcoma virus, baboon endogenous type C virus, and putative human type C viruses [HL23V isolated from blood cells of a patient with acute myelogenous leukemia (HL23) and HEL-12V from human embryonic diploid cells (CIH-32)]. Two sera also reacted with C57BL/6 mouse leukemia induced by Friend virus. These results were obtained by indirect immunoelectron microscopy with various virus-producing cells and by absorption tests using as targets gibbon lymphosarcoma cells that release gibbon ape leukemia virus. In a previous report, the presence of natural antibody in sera from healthy gibbon apes was demonstrated. When the specificities of the human and gibbon natural antibodies were compared, the human natural antibody reacted with two nonproducing culture cell lines of human lymphocytic leukemia (CEM-A and MOLT) and with human embryonic diploid (CIH-1(V-) cells [which became type C virus-producing CIH-32(V+) cells after many passages], but did not react with normal gibbon spleen monolayer cells. In contrast, gibbon natural antibody showed no reaction with CEM-A, MOLT, and CIH-1(V-) cells but reacted with gibbon spleen monolayer cells. Neither human nor gibbon natural antibody that was reactive with gibbon ape leukemia virus crossreacted with feline leukemia virus and mouse wild-type AKR leukemia virus. The gibbon lymphosarcoma cells releasing gibbon ape leukemia virus were used in a screening study of sera from healthy humans. Out of 72 sera screened by indirect immunoelectron microscopy using this system, 55 were positive (76%), i.e., 26 out of 35 males (74%) and 29 out of 37 females (78%). The highest incidence of antibody production was in 1- to 10-year-olds and 31- to 40-year-olds, with the adults exhibiting higher levels. Differences in incidence of natural antibody were not found to be sex-linked. These findings suggest that type C RNA viruses related to the gibbon ape leukemia virus and simian (woolly monkey) sarcoma virus family as well as the baboon endogenous type C virus family may be widespread in humans.

Animals↗

Endogenous N-losses in broilers estimated by a [15N]-isotope dilution technique: effect of dietary fat type and xylanase addition.

Male broilers were given a low protein diet (15.5% CP) spiked with [15N]H4HCO3 from day 12 to day 18 of age to label the endogenous N-constituents. Experimental diets were subsequently fed from day 19 to day 24 of age and consisted of a rye based diet (56% dietary inclusion) which contained either 10% soya oil (S) or 10% beef tallow (T), each of which was either unsupplemented (-) or supplemented (+) with a xylanase containing enzyme preparation (2700 IU/kg at pH 5.3). [15N]-atom percent excess (APE) of excreta, faeces and urine were monitored on a daily basis during both experimental periods. Furthermore, APE was measured in various tissues at the end of the experiment. The APE of urine on the last day of the experiment was between the APE of the pancreas and that of the jejunal tissue, an observation which supported the usefulness of using urinary APE as an indicator for the endogenous N-pool. Endogenous N-proportions were estimated by an isotope dilution technique at the end of the experiment by examination of the ratio of APE in faeces and urine. The endogenous N-proportion in the faeces was greatest in birds receiving the T(-) diet. The proportions were 0.321, 0.319, 0.451 and 0.289 in S(-), S(+), T(-) and T(+) fed groups, respectively. Xylanase addition reduced endogenous N-proportion, a factor which was used to correct apparent crude protein digestibility (85.6, 86.2, 84.3 and 88.5% in S(-), S(+), T(-) and T(+) fed birds, respectively) for endogenous losses resulting in almost equal true digestibilities of crude protein for all treatments (90.3, 90.6, 90.4 and 91.5%). The amounts of endogenous N in faces were estimated to be 87, 69, 244 and 81 mg per day per kg0.67 body weight in S(-), S(+), T(-) and T(+) fed birds, respectively. It was concluded that xylanase supplementation of a rye based broiler diet does not change endogenous N-secretions when the supplemental fat is soya oil. However, addition of tallow rather than soya oil increased these N-losses significantly, an effect which was reversed by xylanase addition.

Animal Feed↗

Human-specific amino acid changes found in 103 protein-coding genes.

We humans have many characteristics that are different from those of the great apes. These human-specific characters must have arisen through mutations accumulated in the genome of our direct ancestor after the divergence of the last common ancestor with chimpanzee. Gene trees of human and great apes are necessary for extracting these human-specific genetic changes. We conducted a systematic analysis of 103 protein-coding genes for human, chimpanzee, gorilla, and orangutan. Nucleotide sequences for 18 genes were newly determined for this study, and those for the remaining genes were retrieved from the DDBJ/EMBL/GenBank database. The total number of amino acid changes in the human lineage was 147 for 26,199 codons (0.56%). The total number of amino acid changes in the human genome was, thus, estimated to be about 80,000. We applied the acceleration index test and Fisher's synonymous/nonsynonymous exact test for each gene tree to detect any human-specific enhancement of amino acid changes compared with ape branches. Six and two genes were shown to have significantly higher nonsynonymous changes at the human lineage from the acceleration index and exact tests, respectively. We also compared the distribution of the differences of the nonsynonymous substitutions on the human lineage and those on the great ape lineage. Two genes were more conserved in the ape lineage, whereas one gene was more conserved in the human lineage. These results suggest that a small proportion of protein-coding genes started to evolve differently in the human lineage after it diverged from the ape lineage.

Animals↗

3'-phosphodiesterase activity of human apurinic/apyrimidinic endonuclease at DNA double-strand break ends.

In order to assess the possible role of human apurinic/apyrimidinic endonuclease (Ape) in double-strand break repair, the substrate specificity of this enzyme was investigated using short DNA duplexes and partial duplexes, each having a single 3'-phosphoglycolate terminus. Phosphoglycolate removal by Ape was detected as a shift in mobility of 5'-end-labeled DNA strands on polyacrylamide sequencing gels, and was quantified by phosphorimaging. Recombinant Ape efficiently removed phosphoglycolates from the 3'-terminus of an internal 1 base gap in a 38mer duplex, but acted more slowly on 3'-phosphoglycolates at a 19 base-recessed 3'-terminus, at an internal nick with no missing bases, and at a double-strand break end with either blunt or 2 base-recessed 3'-termini. There was no detectable activity of Ape toward 3'-phosphoglycolates on 1 or 2 base protruding single-stranded 3'-overhangs. The results suggest that both a single-base internal gap, and duplex DNA on each side of the gap are important binding/recognition determinants for Ape. While Ape may play a role in repair of terminally blocked double-strand breaks, there must also be additional factors involved in removal of at least some damaged 3'-termini, particularly those on 3'-overhangs.

Base Sequence↗

Comparison of quality of life in patients undergoing abdominoperineal extirpation or anterior resection for rectal cancer.

OBJECTIVE: To evaluate the quality of life (QoL) in patients undergoing anterior resection (AR) or abdominoperineal extirpation (APE) for rectal cancer in a sample of patients recruited from a field trial. SUMMARY BACKGROUND DATA: Abdominoperineal resection has been reported to put patients at higher risk of disruption to QoL than sphincter-preserving surgery. METHODS: Fifty patients treated with AR and 23 patients treated with APE were prospectively followed up. All patients were treated in curative attempt and were disease-free throughout the study. QoL was assessed before surgery and 6 to 9 and 12 to 15 months after surgery. RESULTS: Multivariate analysis of variance and subsequent post hoc comparisons revealed a main effect for time (role function, emotional function, body image, future perspective, and micturition-related problems) and group in favor of APE (sleeping problems, constipation, diarrhea), and a time-by-group interaction (role function). No significant results were obtained for the remaining scores, but patients undergoing APE consistently had more favorable QoL scores than those undergoing AR. Multivariate analysis and post hoc comparisons revealed a particularly poor QoL for patients undergoing low AR. They had a significantly lower total QoL, role function, social function, body image, and future perspective, and more gastrointestinal and defecation-related symptoms than patients undergoing high AR. CONCLUSION: Patients undergoing APE do not have a poorer QoL than patients undergoing AR. Patients undergoing low AR have a lower QoL than those undergoing APE. Attention should be paid to QoL concerns expressed by patients undergoing low AR.

Abdomen↗

Autologous plasmin for pharmacologic vitreolysis prepared 1 hour before surgery.

PURPOSE: To evaluate the safety and efficacy of intravitreal injection of autologous plasmin enzyme (APE) in inducing a posterior vitreous detachment (PVD). APE was obtained by a modified method 1 hour before surgery. METHODS: APE was obtained by centrifugation of autologous whole blood from the patients 1 hour before surgery and by incubation with streptokinase. APE was injected in the vitreous cavity 20 minutes before surgery. This procedure was applied for 20 patients who underwent vitrectomy for macular pucker, macular hole, and macular edema due to vitreoretinal traction. The status of PVD was graded intraoperatively. The plasmin concentration obtained by this method and the plasminogen titer of the plasma from each patient were compared. Activated partial thromboplastin time was controlled before surgery and 1, 6, 12, and 24 hours after surgery to evaluate alteration of the coagulation due to systemic absorption of the plasmin. RESULTS: Preparation of APE was easy and took on average 45 minutes for all patients. A PVD with an evident Weiss ring was observed during surgery in 17 eyes (85%). The average plasmin activity was 0.26 IU/0.2 mL activated plasma. The partial thromboplastin time did not show any alteration in any of the patients with respect to preoperative values. CONCLUSION: APE obtained by this method can lead to a PVD and facilitate the complete remotion of vitreous cortex. The intravitreal injection does not lead to alteration of systemic blood coagulation.

Aged↗

Antiphosphatidylethanolamine antibodies in recurrent early pregnancy loss and mid-to-late pregnancy loss.

AIM: Associations have been reported between antiphospholipid antibodies (aPL), mainly anticardiolipin antibodies (aCL) and/or the lupus anticoagulant, and recurrent pregnancy losses (RPL). However, relatively few studies describing antiphosphatidylethanolamine antibodies (aPE) have been reported. We describe the prevalence of aPL to both cardiolipin and phosphatidylethanolamine in patients with RPL. METHODS: Patients with recurrent early pregnancy losses (n = 145) and mid-to-late pregnancy loss(es) (n = 26) were screened for aPE and aCL. RESULTS: In patients with recurrent early pregnancy losses, prevalence of immunoglobulin G (IgG) aPE (17.9%, P = 0.001) and immunoglobulin M (IgM) aPE (12.4%, P = 0.01) was significantly higher than in the control group. In patients with mid-to-late pregnancy loss(es), prevalence of IgM aPE (19.2%, P = 0.008) and IgG aCL (23.1%, P = 0.02) was significantly higher than in the control group. CONCLUSION: Our data suggest that aPE may be a risk factor in patients with mid-to-late pregnancy loss(es) as well as recurrent early pregnancy losses.

Abortion, Habitual↗

Structure-activity relationships of new analogues of arecaidine propargyl ester at muscarinic M1 and M2 receptor subtypes.

1. The potency of arecaidine propargyl ester (APE) and of several analogues containing a modified ester side chain has been assessed at M1 and M2 muscarinic receptor subtypes. APE was shown to act as a potent agonist at ganglionic M1 receptors in the pithed rat, at M2 receptors in guinea-pig isolated atria (-log EC50 = 8.22) and ileum (-log EC50 = 7.77). 2. The arecaidine 2-butynyl and 2-pentynyl esters were approximately equipotent with APE at M1 and M2 receptors, whereas the 2-hexynyl derivative was found to be less potent than APE in atria (-log EC50 = 6.80) and ileum (-log EC50 = 6.70) by about one order of magnitude. The 2-heptynyl and 3-phenyl propargyl esters exhibited no agonist actions in atria and ileum. 3. Shifting the triple bond from the 2 to the 3 position and introducing a bulky group at position 1 of the ester side chain of APE and analogues resulted in competitive antagonists (pA2 ranging from 4.9 to 7.3). 4. APE and its 2-butynyl analogue showed some agonistic selectivity for cardiac M2 receptors (potency ratio, ileum/atria = 2.8 and 4.6 respectively). All antagonists in this series of compounds were not selective in terms of affinity since their pA2 values at cardiac and ileal M2 receptors were similar (potency ratios, ileum/atria = 0.4 to 1.2).

Animals↗

Defining the outcome measures for out-of-hospital trials in acute pulmonary edema.

OBJECTIVE: Comparing studies of the effectiveness of out-of-hospital interventions in acute pulmonary edema (APE) is difficult due to the diversity of outcome measures used in the literature. The objective of this study was to define a set of clinically relevant outcome measures for future out-of-hospital trials in APE. METHODS: A Medline search and hand-search of bibliographies was undertaken to develop a list of APE outcome measures. A survey was mailed to a sample of 227 Canadian emergency physicians using the Dillman methodology, requesting that respondents select clinically relevant outcome measures from this list and rank them by importance. A selection frequency of >or=70% and a median ranking score were used to determine relevant outcome measures. RESULTS: The Medline and bibliography search identified 21 APE outcome measures. The survey response rate was 71%. Outcome measures selected most frequently were heart rate, respiratory rate, respiratory distress scale, subjective dyspnea scale, out-of-hospital intubation, emergency department (ED) intubation, survival to discharge, and out-of-hospital mortality. The median ranking score identified a similar set of measures: heart rate, respiratory rate, respiratory distress scale, subjective dyspnea scale, out-of-hospital intubation rate, and ED intubation rate. There was no significant difference in outcome selection between physicians who worked in communities with and without advanced out-of-hospital care. CONCLUSIONS: Clinically relevant out-of-hospital APE outcome measures were identified and endorsed by a representative survey of Canadian emergency physicians. Clinicians appear to favor short-term and non-mortality outcomes for out-of-hospital interventions. The use of this set of APE outcome measures may improve the design and comparability of future out-of-hospital trials.

Emergency Medical Services↗

Pathogenic role of endothelin 1 in hemodynamic dysfunction in experimental acute pulmonary thromboembolism.

The plasma endothelin-1 (ET-1) level is elevated in patients with acute pulmonary thromboembolism (APE). Whether ET-1 is a pathogenic mediator or a simple marker of APE is not known. We investigated the role of ET-1 in hemodynamic dysfunction in APE through evaluating the effects of ET(A) receptor antagonist in an experimental APE model. We also examined ET-1 expression in embolized lungs. In a canine autologous blood clot pulmonary embolism model, ET(A) receptor antagonist ZD2574 (10 mg/kg, intravenous; ZD2574 group; n = 6) or vehicle (control group; n = 5) was administered. Hemodynamic and gas exchange parameters and plasma levels of ET-1 were serially measured. Prepro-ET-1 mRNA expression and the distribution of ET-1 peptide in lung tissues were also examined. With ZD2574 pulmonary arterial pressure and pulmonary vascular resistance significantly decreased, and were lower compared with the control group. The decrease in cardiac output was also less in the ZD2574 group. Plasma ET-1 levels increased after embolization. Prepro-ET-1 mRNA expression increased in embolized lungs and ET-1 peptide expression also increased in embolized lungs, particularly in the muscular pulmonary arteries, compared with normal lungs. These findings suggest that ET-1 partially contributes to hemodynamic derangements of APE, and that ET(A) receptor antagonists might constitute a useful therapeutic tool for APE.

Acute Disease↗

Total sequence decomposition distinguishes functional modules, "molegos" in apurinic/apyrimidinic endonucleases.

BACKGROUND: Total sequence decomposition, using the web-based MASIA tool, identifies areas of conservation in aligned protein sequences. By structurally annotating these motifs, the sequence can be parsed into individual building blocks, molecular legos ("molegos"), that can eventually be related to function. Here, the approach is applied to the apurinic/apyrimidinic endonuclease (APE) DNA repair proteins, essential enzymes that have been highly conserved throughout evolution. The APEs, DNase-1 and inositol 5'-polyphosphate phosphatases (IPP) form a superfamily that catalyze metal ion based phosphorolysis, but recognize different substrates. RESULTS: MASIA decomposition of APE yielded 12 sequence motifs, 10 of which are also structurally conserved within the family and are designated as molegos. The 12 motifs include all the residues known to be essential for DNA cleavage by APE. Five of these molegos are sequentially and structurally conserved in DNase-1 and the IPP family. Correcting the sequence alignment to match the residues at the ends of two of the molegos that are absolutely conserved in each of the three families greatly improved the local structural alignment of APEs, DNase-1 and synaptojanin. Comparing substrate/product binding of molegos common to DNase-1 showed that those distinctive for APEs are not directly involved in cleavage, but establish protein-DNA interactions 3' to the abasic site. These additional bonds enhance both specific binding to damaged DNA and the processivity of APE1. CONCLUSION: A modular approach can improve structurally predictive alignments of homologous proteins with low sequence identity and reveal residues peripheral to the traditional "active site" that control the specificity of enzymatic activity.

Amino Acid Motifs↗