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J Mendes

Publications and source records attributed to J Mendes.

At least 19 recordsLinked to original sources
PubMed2004

Mite allergen levels and acarologic analysis in house dust samples in Uberaba, Brazil.

Mite allergen exposure has been widely related to sensitization and development of allergic diseases. This study intended to evaluate the degree of allergen exposure in Uberaba, Brazil, through the measurements of Der f 1 and Der p 1 allergen levels associated with the acarologic analysis in house dust samples. A total of 240 dust samples were collected from 60 houses through vacuuming sofas and bedding, during the months of March and July 2000. Indoor temperature and relative humidity were also measured. Mites were counted and identified under light microscopy and allergen levels were measured by two-site monoclonal antibody ELISAs. The major mite family was Pyroglyphidae (39.4%), having D. pteronyssinus as the most frequent species (15.6%), followed by D. farinae (12.3%) and E. maynei (7.9%). The family Glycyphagidae was less commonly found (4.8%), with Blomia tropicalis (4.4%) as its majoritary member. The highest levels of Der f 1 and Der p 1 allergens were found in bedding samples in March (31.7 and 0.9 microg/g of dust, respectively), with Der f 1 levels significantly higher than Der p 1 (p < 0.0001). There was a significant positive correlation between the mite number and allergen levels. These results indicate that Dermatophagoides sp are the most frequent mites in our region followed by E. maynei. Therefore, the knowledge of the local mite fauna would improve the means of investigating the association between allergen exposure and sensitization, allowing the addition of new mite extracts in diagnostic tests.

Air Pollution, Indoor↗
PubMed2002

Peyronie's disease: surgical correction of 40 patients with relaxing incision and duramater graft.

OBJECTIVE: Assessment of a long-term follow-up of a previously described technique to correct the curvature of the penis in Peyronie's disease. MATERIAL AND METHODS: Forty patients (43-69 years old) were operated between 1989 and 1999. The eligibility criteria for surgery were: lesion stabilised during, at least, one year; angulation of 40 degrees or more; and difficult or impossible sexual intercourse. Moreover, the patients had to have good libido, maintained erections and interested sexual partners. The operative technique consisted in making an I-shaped relaxing incision in the zone of higher angulation of the tunica albuginea and the defect created in the tunica was covered with a human duramater graft. RESULTS: The follow-up varied from 1 to 6 years (70%, > or =2 years). Cosmetic results: 95% good, 5% satisfactory (not needing further correction). Functional results: 85% good, 15% erectile dysfunction (10% moderate and 5% severe). COMPLICATIONS: one recurrence, one partial necrosis of the foreskin, nine decreases in the sensibility of the glans (eight recovered after 4-12 months post-operatively). CONCLUSIONS: The I-shaped relaxing incision and duramater graft procedure had a satisfactory success rate in the correction of severe Peyronie's disease. The utilisation of ready to use material (homo or hetero graft) avoids additional surgery, reduces the operative time and possible complications. Like in other procedures, the major problem of this surgery is the erectile dysfunction rate.

Adult↗
PubMed2001

Incorporating knowledge-based biases into an energy-based side-chain modeling method: application to comparative modeling of protein structure.

The performance of the self-consistent mean field theory (SCMFT) method for side-chain modeling, employing rotamer energies calculated with the flexible rotamer model (FRM), is evaluated in the context of comparative modeling of protein structure. Predictions were carried out on a test set of 56 model backbones of varying accuracy, to allow side-chain prediction accuracy to be analyzed as a function of backbone accuracy. A progressive decrease in the accuracy of prediction was observed as backbone accuracy decreased. However, even for very low backbone accuracy, prediction was substantially higher than random, indicating that the FRM can, in part, compensate for the errors in the modeled tertiary environment. It was also investigated whether the introduction in the FRM-SCMFT method of knowledge-based biases, derived from a backbone-dependent rotamer library, could enhance its performance. A bias derived from the backbone-dependent rotamer conformations alone did not improve prediction accuracy. However, a bias derived from the backbone-dependent rotamer probabilities improved prediction accuracy considerably. This bias was incorporated through two different strategies. In one (the indirect strategy), rotamer probabilities were used to reject unlikely rotamers a priori, thus restricting prediction by FRM-SCMFT to a subset containing only the most probable rotamers in the library. In the other (the direct strategy), rotamer energies were transformed into pseudo-energies that were added to the average potential energies of the respective rotamers, thereby creating hybrid energy-based/knowledge-based average rotamer energies, which were used by the FRM-SCMFT method for prediction. For all degrees of backbone accuracy, an optimal strength of the knowledge-based bias existed for both strategies for which predictions were more accurate than pure energy-based predictions, and also than pure knowledge-based predictions. Hybrid knowledge-based/energy-based methods were obtained from both strategies and compared with the SCWRL method, a hybrid method based on the same backbone-dependent rotamer library. The accuracy of the indirect method was approximately the same as that of the SCWRL method, but that of the direct method was significantly higher.

Computer Simulation↗
PubMed2001

Implicit solvation in the self-consistent mean field theory method: sidechain modelling and prediction of folding free energies of protein mutants.

The Atomic Solvation Parameter (ASP) model is one of the simplest models of solvation, in which the solvation free energy of a molecule is proportional to the solvent accessible surface area (SAS) of its atoms. However, until now this model had not been incorporated into the Self-Consistent Mean Field Theory (SCMFT) method for modelling sidechain conformations in proteins. The reason for this is that SAS is a many-body quantity and, thus, it is not obvious how to define it within the Mean Field (MF) framework, where multiple copies of each sidechain exist simultaneously. Here, we present a method for incorporating an SAS-based potential, such as the ASP model, into SCMFT. The theory on which the method is based is exact within the MF framework, that is, it does not depend on a pairwise or any other approximation of SAS. Therefore, SAS can be calculated to arbitrary accuracy. The method is computationally very efficient: only 7.6% slower on average than the method without solvation. We applied the method to the prediction of sidechain conformation, using as a test set high-quality solution structures of 11 proteins. Solvation was found to substantially improve the prediction accuracy of well-defined surface sidechains. We also investigated whether the methodology can be applied to prediction of folding free energies of protein mutants, using a set of barnase mutants. For apolar mutants, the modest correlation observed between calculated and observed folding free energies without solvation improved substantially when solvation was included, allowing the prediction of trends in the folding free energies of this type of mutants. For polar mutants, correlation was not significant even with solvation. Several other factors also responsible for the correlation were identified and analysed. From this analysis, future directions for applying and improving the present methodology are discussed.

Algorithms↗
PubMed2000

Evolution of the thyroid hormone-binding protein, transthyretin.

Transthyretin (TTR) belongs to a group of proteins, which includes thyroxine-binding globulin and albumin, that bind to and transport thyroid hormones in the blood. TTR is also indirectly implicated in the carriage of vitamin A through the mediation of retinol-binding protein (RBP). It was first identified in 1942 in human serum and cerebrospinal fluid and was formerly called prealbumin for its ability to migrate faster than serum albumin on electrophoresis of whole plasma. It is a single polypeptide chain of 127 amino acids (14,000 Da) and is present in the plasma as a tetramer of noncovalently bound monomers. The major sites of synthesis of TTR in eutherian mammals, marsupials, and birds are the liver and choroid plexus but in reptiles it is synthesised only in the choroid plexus. The observation that TTR is strongly expressed in the choroid plexus but not in the liver of the stumpy-tailed lizard and the strong conservation of expression in the choroid plexus from reptiles to mammals have been taken as evidence to suggest that extrahepatic synthesis of TTR evolved first. The identification and cloning of TTR from the liver of an amphibian, Rana catesbeiana, and a teleost fish, Sparus aurata, and its absence from the choroid plexus of both species suggest an alternative model for its evolution. Protein modelling studies are presented that demonstrate differences in the electrostatic characteristics of the molecule in human, rat, chicken, and fish, which may explain why, in contrast to TTR from human and rat, TTR from fish and birds preferentially binds triiodo-l-thyronine.

Amino Acid Sequence↗
PubMed2000

Latin american experience with two low-dose oral contraceptives containing 30 microg ethinylestradiol/75 microg gestodene and 20 microg ethinylestradiol/150 microg desogestrel.

The objective of this study was to compare cycle control, efficacy and tolerance of an oral contraceptive containing 20 microg ethinylestradiol and 150 microg desogestrel with a preparation containing 30 microg ethinylestradiol combined with 75 microg gestodene. This study involved 342 women and 4104 cycles use in Argentina, Brazil, Chile, and Mexico. Contraceptive efficacy was good with both formulations. Two pregnancies occurred in the desogestrel group but were not due to method failure. With respect to cycle control, the incidence of intermenstrual bleeding was higher during the first 3 cycles in the desogestrel group; it was significant (p <0.01) during the first 3 days of the cycle for a normal or heavy bleeding only in the Mexican group. Amenorrhea was not reported for any group, but the incidence of dysmenorrhea was significantly higher (p <0.01) in the Brazilian desogestrel group (13.8%) and was significantly lower (p <0.01) in the Mexican gestodene group (8.5%). Adverse events were similar in all the countries with headache, breast tension, and nausea, the most frequently reported symptoms. The range of mean increase in body weight varied from 0.2 kg in the Argentine group to 2.6 kg in the Chilean group (95% confidence limit, +/- 2.51) in the gestodene group, and 0.2 kg in the Argentine group to 2.5 kg in Brazilian group (95% confidence limit, +/- 2.36) in the desogestrel group. Fifteen women discontinued because of headache, but there were no significant differences between the groups regarding discontinuation for this and other medical or non-medical reasons. Both oral contraceptive preparations are reliable and well tolerated, and both have favorable effects on control cycle.

Adolescent↗
PubMed1999

Improved modeling of side-chains in proteins with rotamer-based methods: a flexible rotamer model.

Side-chain modeling has a widespread application in many current methods for protein tertiary structure determination, prediction, and design. Of the existing side-chain modeling methods, rotamer-based methods are the fastest and most efficient. Classically, a rotamer is conceived as a single, rigid conformation of an amino acid sidechain. Here, we present a flexible rotamer model in which a rotamer is a continuous ensemble of conformations that cluster around the classic rigid rotamer. We have developed a thermodynamically based method for calculating effective energies for the flexible rotamer. These energies have a one-to-one correspondence with the potential energies of the rigid rotamer. Therefore, the flexible rotamer model is completely general and may be used with any rotamer-based method in substitution of the rigid rotamer model. We have compared the performance of the flexible and rigid rotamer models with one side-chain modeling method in particular (the self-consistent mean field theory method) on a set of 20 high quality crystallographic protein structures. For the flexible rotamer model, we obtained average predictions of 85.8% for chi1, 76.5% for chi1+2 and 1.34 A for root-mean-square deviation (RMSD); the corresponding values for core residues were 93.0%, 87.7% and 0.70 A, respectively. These values represent improvements of 7.3% for chi1, 8.1% for chi1+2 and 0.23 A for RMSD over the predictions obtained with the rigid rotamer model under otherwise identical conditions; the corresponding improvements for core residues were 6.9%, 10.5% and 0.43 A, respectively. We found that the predictions obtained with the flexible rotamer model were also significantly better than those obtained for the same set of proteins with another state-of-the-art side-chain placement method in the literature, especially for core residues. The flexible rotamer model represents a considerable improvement over the classic rigid rotamer model. It can, therefore, be used with considerable advantage in all rotamer-based methods commonly applied to protein tertiary structure determination, prediction, and design and also in predictions of free energies in mutational studies.

Amino Acids↗
PubMed1999

The caa3 terminal oxidase of the thermohalophilic bacterium Rhodothermus marinus: a HiPIP:oxygen oxidoreductase lacking the key glutamate of the D-channel.

The respiratory chain of the thermohalophilic bacterium Rhodothermus marinus contains a novel complex III and a high potential iron-sulfur protein (HiPIP) as the main electron shuttle (Pereira et al., Biochemistry 38 (1999) 1268-1275 and 1276-1283). In this paper, one of the terminal oxidases expressed in this bacterium is extensively characterised. It is a caa3-type oxidase, isolated with four subunits (apparent molecular masses of 42, 19 and 15 kDa and a C-haem containing subunit of 35 kDa), which has haems of the A(s) type. This oxidase is capable of using TMPD and horse heart cytochrome c as substrates, but has a higher turnover with HiPIP, being the first example of a HiPIP:oxygen oxidoreductase. The oxidase has unusually low reduction potentials of 260 (haem C), 255 (haem A) and 180 mV (haem A3). Subunit I of R. marinus caa3 oxidase has an overall significant homology with the subunits I of the COX type oxidases, namely the metal binding sites and most residues considered to be functionally important for proton uptake and pumping (K- and D-channels). However, a major difference is present: the putative essential glutamate (E278 in Paraccocus denitrificans) of the D-channel is missing in the R. marinus oxidase. Homology modelling of the R. marinus oxidase shows that the phenol group of a tyrosine residue may occupy a similar spatial position as the glutamate carboxyl, in relation to the binuclear centre. Moreover, sequence comparisons reveal that several enzymes lacking that glutamate have a conserved substitution pattern in helix VI: -YSHPXV- instead of -XGHPEV-. These observations are discussed in terms of the mechanisms for proton uptake and it is suggested that, in these enzymes, tyrosine may play the role of the glutamate in the proton channel.

Amino Acid Sequence↗
PubMed1999

An iterative structure-assisted approach to sequence alignment and comparative modeling.

Correct alignment of the sequence of a target protein with those of homologues of known three-dimensional structure is a key step in comparative modeling. Usually an iterative approach that takes account of the local and overall structural features is required. We describe such an approach that exploits databases of structural alignments of homologous proteins (HOMSTRAD, http:/(/)www-cryst.bioc.cam.ac.uk/ approximately homstrad) and protein superfamilies (CAMPASS, http:/(/)www-cryst.bioc.cam.ac.uk/ approximately campass), in which structure-based alignments are analyzed and formatted with the program JOY (http:/(/)www-cryst.bioc.cam.ac.uk/ approximately joy) to reveal conserved local structural features. The databases facilitate the recognition of a family or superfamily, they assist in the selection of useful parent structures, they are helpful in alignment of the target sequences with the parent set, and are useful for deriving relationships that can be used in validating models. In the iterative approach, a model is constructed on the basis of the proposed sequence alignment and this is then reexpressed in the JOY format and realigned with the parent set. This is repeated until the model and sequence alignment is optimized. We examine the case for comparison and use of multiple structures of family members, rather than a single parent structure. We use the targets attempted by our group in CASP3 to assess the value of such procedures.

Algorithms↗
PubMed1999

Improvement of side-chain modeling in proteins with the self-consistent mean field theory method based on an analysis of the factors influencing prediction.

With the objective of improving side-chain conformation prediction, we have analyzed the influence of various factors on prediction by the Self-Consistent Mean Field Theory method, applied to a set of high resolution x-ray protein structure models. These factors may be classed as variations in the mean field optimization protocol, variations in the potential energy function, and variations in rotamer library completeness. We have developed an optimization protocol that consistently reached lower mean field conformational free energies than two other protocols. This protocol led to an important improvement in prediction. We observed a major improvement in prediction with two more detailed van der Waals parameter sets, which we found to be due mainly to the introduction of scaling of 1-4 interactions. In a comparison of two knowledge-based rotamer libraries of considerably different size, we observed an unexpected decrease in prediction with an increase in library completeness. However, when we introduced a torsion potential term in the potential energy function, we found an important increase in average prediction and in the prediction of almost all residue types with a more complete rotamer set. The two knowledge-based rotamer libraries now became equivalent in terms of average prediction. The results we obtained in an analysis of the effect of the introduction of an additional electrostatic term in the potential energy function were largely inconclusive. However, we found a small increase in average prediction for an electrostatic potential term with a fixed dielectric constant of 15. The combined effect of all the factors we analyzed in this study resulted in average prediction accuracies of 79.9% for X1, 68.1% for X1 + 2, and 1.590 A for global rms deviation (RMSD); the corresponding values for core residues were 88.2%, 78.6%, and 1.171 A. These values represent improvements in average prediction of 6.5% for X1, 9.1% for X1 + 2, and 0.163 A for global RMSD over the original conditions; the corresponding improvements in the core were 5.9%, 9.0%, and 0.180 A, respectively.

Amino Acids↗
PubMed1999

Diapause, pupation sites and parasitism of the horn fly, Haematobia irritans, in south-eastern Brazil.

In order to verify the occurrence of diapause, preference for pupation sites and hymenopteran parasitism, the pupae of the horn fly, Haematobia irritans (Diptera: Muscidae), were collected from undisturbed cattle dung pats in pastures, and adults of the fly were sampled from cattle in São Paulo State, south-eastern Brazil, from April 1993 to July 1994. Diapause was verified in 7.7% of pupae sampled from pastures in June and July of 1993 and in 9.9% of those sampled in May, June and July of 1994 (overall rate of 9.1%). Approximately 8.3% of the pupae were parasitized by microhymenopterans, mostly Spalangia nigroaenea and S. cameroni (Hymenoptera: Pteromalidae). Horn fly pupae were found almost exclusively inside the pat or in the soil immediately beneath and adjacent to it, and very few were collected elsewhere. Pupa mortality was 54.4% and did not change significantly during the year, but mortality was greater among pupae collected in pastures when compared to those obtained from experimental pats, lacking natural enemies.

Animals↗
PubMed1999

[Comparative study of Toronto SPV prosthesis and bileaflet mechanical aortic prosthesis with 2D-Doppler echocardiography].

AIM: The aim of this study was to make a noninvasive comparison, by means of Doppler echocardiography, of the hemodynamic performance of biological stentless xenografts and mechanical bileaflet (MB) prostheses (P) in aortic position. METHODS: We studied 20 patients (pts) with normofunctional (nf) aortic Toronto (T) Stentless Porcine valves (SPV)--Group I--8 males, aged 69 +/- 12 years, 32 +/- 9 months after surgery, and 30 pts with nf MB aortic P (Carbomedics or St. Jude Medical)--Group II--17 males (p = NS vs G I), aged 61 +/- 12 years (p < 0.01 vs G I), 30 +/- 12 months after implantation (p = NS vs G I). Both groups were comparable with regard to body surface area and surgical indication. P diameters ranged from 21 to 25 mm (G I: 22.9 +/- 1.7; G II: 22.8 +/- 1.7 - p = NS), the number of pts with the same P diameter in each group being similar. We analysed, at rest: aortic orifice diameter (AoOd - cm), maximal (GMax) and mean (GMean) transprosthesic pressure gradients, P functional area (PFA) and P resistance (PRes). Gradients (mm Hg) were calculated by means of the Bernoulli equation, PFA (cm2) by means of the continuity equation and PRes (dynes.s.cm-5) as 1333 x Gmean x SEP/SV (SEP = systolic ejection period; SV = stroke volume). RESULTS: AoOd (G I vs G II): P 21--1.78 +/- 0.04 vs 2.00 +/- 0.10 (p < 0.001); P 23--1.91 +/- 0.10 vs 2.19 +/- 0.10 (p = 0.01); P 25--2.22 +/- 0.24 vs 2.29 +/- 0.19 (NS). Doppler parameters: [table: see text] CONCLUSIONS: P T SPV show better hemodynamic performance when compared to P MB with the same diameter, in aortic position. In addition, our results suggest that P T SPV allow the use of larger valve sizes for the same aortic orifice diameter.

Aged↗
PubMed1998

Molecular dynamics simulation of cytochrome c3: studying the reduction processes using free energy calculations.

The tetraheme cytochrome c3 from Desulfovibrio vulgaris Hildenborough is studied using molecular dynamics simulation studies in explicit solvent. The high heme content of the protein, which has its core almost entirely made up of c-type heme, presents specific problems in the simulation. Instability in the structure is observed in long simulations above 1 ns, something that does not occur in a monoheme cytochrome, suggesting problems in heme parametrization. Given these stability problems, a partially restrained model, which avoids destruction of the structure, was created with the objective of performing free energy calculations of heme reduction, studies that require long simulations. With this model, the free energy of reduction of each individual heme was calculated. A correction in the long-range electrostatic interactions of charge groups belonging to the redox centers had to be made in order to make the system physically meaningful. Correlation is obtained between the calculated free energies and the experimental data for three of four hemes. However, the relative scale of the calculated energies is different from the scale of the experimental free energies. Reasons for this are discussed. In addition to the free energy calculations, this model allows the study of conformational changes upon reduction. Even if the precise details of the structural changes that take place in this system upon individual heme reduction are probably out of the reach of this study, it appears that these structural changes are small, similarly to what is observed for other redox proteins. This does not mean that their effect is minor, and one example is the conformational change observed in propionate D from heme I when heme II becomes reduced. A motion of this kind could be the basis of the experimentally observed cooperativity effects between heme reduction, namely positive cooperativity.

Biophysical Phenomena↗
PubMed1998

[Congenital bronchopulmonary cystic disease].

From September 1989 to December 1994, 12 children with congenital bronchopulmonary cystic disease were operated in the Department of Pediatric Surgery, Santa Maria Hospital. Four cases of bronchogenic cyst (BC), 2 of cystic adenomatoid malformation (CAM), 3 of pulmonary sequestration (PS), and 3 of congenital lobar emphysema (CLE) were found. Age ranged between 9 days and 10 years without predominance of gender. Seven were younger than one year of age, of which 5 were less than 6 months old at the time of surgery. The CAT scan was most useful in the diagnosis and follow-up of these children. The review of these cases helped evaluate our experience in the diagnosis and surgical approach for these congenital malformations. Successful treatment depends on early diagnosis and the planning of therapeutic priorities, based on the embryological and pathophysiological knowledge at these anomalies. The embryological mechanisms implicated in the development are discussed and the results of surgical treatment are presented.

Bronchogenic Cyst↗
PubMed1997

[Mitral valve insufficiency caused by tendinous cord rupture and mitral valve aneurysm. Significance of the echocardiographic study].

Mitral valve aneurysm is a rare complication that may occur in a myxomatous valve. We report the case of a 73 year old male patient with severe mitral regurgitation and heart failure-class IV NYHA. Echocardiography showed perforation of an aneurysm of the anterior leaflet of the mitral valve associated with rupture of tendinous cords of the posterior mitral leaflet. Diagnosis was made by transthoracic echocardiography and confirmed by transesophageal echocardiography. The patient was urgently operated with success and a mitral valve prosthesis was implanted.

Aged↗
PubMed1996

Paramagnetic NMR analysis of the seven-iron ferredoxin from the hyperthermoacidophilic archaeon Desulfurolobus ambivalens reveals structural similarity to other dicluster ferredoxins.

The seven-iron ferredoxin from the hyperthermophilic archaeon Desulfurolobus ambivalens has been investigated by one-dimensional and two-dimensional 1H-NMR in its oxidized and dithionite-reduced states. All iron atoms of both the three-iron and the four-iron cluster are bound to cysteine residues whose hyperfine-shifted resonances were characterized. The pattern of these resonances is similar to those from three-iron, four-iron and eight-iron ferredoxins previously described in the literature, but the four-iron cluster has a shift pattern different from that in other seven-iron proteins. A second set of hyperfine-shifted resonances clearly indicates sample heterogeneity, which possibly involves the four-iron cluster. The observation of interresidue NOEs between two different cysteine residues proves the existence of close spatial proximity of the two clusters in D. ambivalens ferredoxin and therefore indicates structural homology to other dicluster ferredoxins. Moreover, this feature is crucial for the sequence-specific assignment of the hyperfine-shifted resonances. The C alpha-C beta-S-Fe dihedral angles of the cysteine residues coordinating the four-iron cluster could be estimated, and the electronic structure of the three-iron cluster is discussed.

Ferredoxins↗
PubMed1995

A seven-iron ferredoxin from the thermoacidophilic archaeon Desulfurolobus ambivalens.

A seven-iron ferredoxin was isolated from aerobically grown cells of the hyperthermoacidophilic archaeon Desulfurolobus ambivalens (DSM 3772). The protein is monomeric, with an apparent molecular mass of 15 kDa and contains 7 iron atoms/molecule. The N-terminal sequence shows a large similarity (70% identity) with that of the ferredoxin isolated from the archaeon Sulfolobus acidocaldarius. The EPR characteristics in both the native (oxidized) and dithionite-reduced states of this protein allowed an unequivocal identification of a [3Fe-4S]1+/0 center, with a reduction potential of -270 +/- 20 mV, at pH 7.5. The protein also contains a [4Fe-4S]2+/1+ center with a very low reduction potential (Eo = -540 mV, pH 7.0), which yields a rhombic EPR spectrum upon reduction with sodium dithionite at high pH. The reduction potentials of both centers are slightly pH dependent between pH 6 and 9. The [3Fe-4S] ferredoxin center is able to accept electrons from pyruvate oxidase and NADH oxidase isolated from D. ambivalens. This ferredoxin is present in large amounts (at least 130 mg/kg wet cells), which allowed the unequivocal observation of oxidized [3Fe-4S] clusters in intact D. ambivalens cells.

Amino Acid Sequence↗