Identifying sagittal otoliths of Mediterranean Sea gobies: variability among phylogenetic lineages.

In this study, we describe and analyse the morphology of the sagitta, the largest otolith, of 25 species of Gobiidae inhabiting the Adriatic and north-western Mediterranean seas. Our goal was to test the usefulness and efficiency of sagittal otoliths for species identification. Our analysis of otolith contours was based on mathematical descriptors called wavelets, which are related to multi-scale decompositions of contours. Two methods of classification were used: an iterative system based on 10 wavelets that searches the Anàlisi de Formes d'Otòlits (AFORO) database and a discriminant method based only on the fifth wavelet. With the exception of paedomorphic species, the results showed that otolith anatomy and morphometry can be used as diagnostic characters distinguishing the three Mediterranean phylogenetic goby lineages (Pomatoschistus or sand-goby lineage, Aphia lineage and Gobius lineage). The main anatomical differences were related to overall shape (square to rhomboid), the development and shape of the postero-dorsal and antero-ventral lobes and the degree of convexity of dorsal and ventral margins. Iterative classifications and discriminant analysis of otolith contour provided very similar results. In both cases, more than 70% of specimens were correctly classified to species and more than 80% to genus. Iterations in the larger AFORO database (including 216 families of teleosts) attained a 100% correct classification at the family level.

Noninvasive Liver Assessment in Adult Patients With Fontan Circulation Using Acoustic Radiation Force Impulse Elastography and Hepatic Magnetic Resonance Imaging.

BACKGROUND: Patients who have undergone the Fontan procedure are at risk of developing hepatic dysfunction. However, broad recommendations regarding liver monitoring are limited. The purpose of this study was to characterize the frequency of liver disease in adult Fontan patients using multimodality imaging (hepatic magnetic resonance imaging [MRI], acoustic radiation force impulse [ARFI] elastography, or hepatic ultrasound). METHODS: In a prospective cross-sectional analysis of adult patients palliated with a Fontan procedure, hepatic MRI, ARFI, and hepatic ultrasound were used to assess for liver disease. The protocol compared (1) varying prevalence of liver disease based on each imaging technique, (2) agreement between different techniques, and (3) association between noninvasive imaging diagnosis of liver disease and clinical variables, including specific liver disease biomarkers. RESULTS: Thirty-seven patients were enrolled. The ARFI results showed high wave propagation velocity in 35 patients (94.6%). All patients had some abnormality in the hepatic MRI. Specifically, 8 patients (21.6%) showed signs of chronic liver disease, 10 patients (27%) had significant liver fibrosis, and 27 patients (73%) had congestion. No correlation was found between liver stiffness measured as propagation velocity and hepatic MRI findings. Only 7 patients had an abnormal hepatic ultrasound study. CONCLUSIONS: There is an inherent liver injury in adult Fontan patients. Signs of liver disease were observed in most patients by both hepatic MRI and ARFI elastography but not by ultrasound imaging. Increased liver stiffness did not identify specific disease patterns from MRI, supporting the need for multimodality imaging to characterize liver disease in Fontan patients.

Otolith shape lends support to the sensory drive hypothesis in rockfishes.

The sensory drive hypothesis proposes that environmental factors affect both signalling dynamics and the evolution of signals and receivers. Sound detection and equilibrium in marine fishes are senses dependent on the sagittae otoliths, whose morphological variability appears intrinsically linked to the environment. The aim of this study was to understand if and which environmental factors could be conditioning the evolution of this sensory structure, therefore lending support to the sensory drive hypothesis. Thus, we analysed the otolith shape of 42 rockfish species (Sebastes spp.) to test the potential associations with the phylogeny, biological (age), ecological (feeding habit and depth distribution) and biogeographical factors. The results showed strong differences in the otolith shapes of some species, noticeably influenced by ecological and biogeographical factors. Moreover, otolith shape was clearly conditioned by phylogeny, but with a strong environmental effect, cautioning about the use of this structure for the systematics of rockfishes or other marine fishes. However, our most relevant finding is that the data supported the sensory drive hypothesis as a force promoting the radiation of the genus Sebastes. This hypothesis holds that adaptive divergence in communication has significant influence relative to other life history traits. It has already been established in Sebastes for visual characters and organs; our results showed that it applies to otolith transformations as well (despite the clear influence of feeding and depth), expanding the scope of the hypothesis to other sensory structures.

X-ray snapshot of HIV-1 protease in action: observation of tetrahedral intermediate and short ionic hydrogen bond SIHB with catalytic aspartate.

Structural snapshots of each step in the catalytic cycle would help development of inhibitors of human immunodeficiency virus type 1 protease (HIV-1 PR) as effective drugs against HIV/AIDS. We report here one snapshot obtained by determining the structure of enzyme-substrate complex under conditions where the catalytic activity of the enzyme is greatly reduced. The 1.76 A crystal structure shows the oligopeptide substrate, AETFYVDGAA, converted in situ into a gem-diol tetrahedral intermediate (TI). The gem-diol intermediate is neutral and one of the hydroxyl oxygens forms a very short hydrogen bond (2.2 A) with the anionic aspartate of the catalytic dyad, which is monoprotonated. Further, there is no hydrogen atom on the outer oxygen of the neutral aspartate. These two observations provide direct evidence that, in the reaction mechanism, hydrogen bonding between catalytic aspartate and scissile carbonyl oxygen facilitates water attack on the scissile carbon atom. Comparison with the structural snapshot of the biproduct complex involving the same substrate reveals the reorganization of the hydrogen bonds at the catalytic center as the enzymatic reaction progresses toward completion. Accumulation of TI in the crystals provides direct evidence that collapse of TI is the rate-limiting step of hydrolysis.

Resistance mechanism revealed by crystal structures of unliganded nelfinavir-resistant HIV-1 protease non-active site mutants N88D and N88S.

Nelfinavir is an inhibitor of HIV-1 protease, and is used for treatment of patients suffering from HIV/AIDS. However, treatment results in drug resistant mutations in HIV-1 protease. N88D and N88S are two such mutations which occur in the non-active site region of the enzyme. We have determined crystal structures of unliganded N88D and N88S mutants of HIV-1 protease to resolution of 1.65A and 1.8A, respectively. These structures refined against synchrotron data lead to R-factors of 0.1859 and 0.1780, respectively. While structural effects of N88D are very subtle, the mutation N88S has caused a significant conformational change in D30, an active site residue crucial for substrate and inhibitor binding.

Structural characterization of the active form of PerR: insights into the metal-induced activation of PerR and Fur proteins for DNA binding.

In Bacillus subtilis, the transcription factor PerR is an iron dependant sensor of H(2)O(2). The sensing mechanism relies on a selective metal catalysed oxidation of two histidine residues of the regulatory site. Here we present the first crystal structure of the active PerR protein in complex with a Mn(2+) ion. In addition, X-ray absorption spectroscopy experiments were performed to characterize the corresponding iron form of the protein. Both studies reveal a penta-coordinate arrangement of the regulatory site that involves three histidines and two aspartates. One of the histidine ligand belongs to the N-terminal domain. Binding of this residue to the regulatory metal allows the protein to adopt a caliper-like conformation suited to DNA binding. Since this histidine is conserved in all PerR and a vast majority of Fur proteins, it is likely that the allosteric switch induced by the regulatory metal is general for this family of metalloregulators.

Upgrade of the CATS sample changer on FIP-BM30A at the ESRF: towards a commercialized standard.

An upgraded version of the sample changer ;CATS' (Cryogenic Automated Transfer System) that was developed on the FIP-BM30A beamline at the ESRF is presented. At present, CATS is installed at SLS (three systems), BESSY (one system), DLS (two systems) and APS (four systems for the LSCAT beamline). It consists mainly of an automated Dewar with an assortment of specific grippers designed to obtain a fast and reliable mounting/dismounting rate without jeopardizing the flexibility of the system. The upgraded system has the ability to manage any sample standard stored in any kind of puck.

X-ray structure of HIV-1 protease in situ product complex.

HIV-1 protease is an effective target for design of different types of drugs against AIDS. HIV-1 protease is also one of the few enzymes that can cleave substrates containing both proline and nonproline residues at the cleavage site. We report here the first structure of HIV-1 protease complexed with the product peptides SQNY and PIV derived by in situ cleavage of the oligopeptide substrate SQNYPIV, within the crystals. In the structure, refined against 2.0-A resolution synchrotron data, a carboxyl oxygen of SQNY is hydrogen-bonded with the N-terminal nitrogen atom of PIV. At the same time, this proline nitrogen atom does not form any hydrogen bond with catalytic aspartates. These two observations suggest that the protonation of scissile nitrogen, during peptide bond cleavage, is by a gem-hydroxyl of the tetrahedral intermediate rather than by a catalytic aspartic acid.

Structure and function of enzymes involved in the biosynthesis of phenylpropanoids.

As a major component of plant specialized metabolism, phenylpropanoid biosynthetic pathways provide anthocyanins for pigmentation, flavonoids such as flavones for protection against UV photodamage, various flavonoid and isoflavonoid inducers of Rhizobium nodulation genes, polymeric lignin for structural support and assorted antimicrobial phytoalexins. As constituents of plant-rich diets and an assortment of herbal medicinal agents, the phenylpropanoids exhibit measurable cancer chemopreventive, antimitotic, estrogenic, antimalarial, antioxidant and antiasthmatic activities. The health benefits of consuming red wine, which contains significant amounts of 3,4',5-trihydroxystilbene (resveratrol) and other phenylpropanoids, highlight the increasing awareness in the medical community and the public at large as to the potential dietary importance of these plant derived compounds. As recently as a decade ago, little was known about the three-dimensional structure of the enzymes involved in these highly branched biosynthetic pathways. Ten years ago, we initiated X-ray crystallographic analyses of key enzymes of this pathway, complemented by biochemical and enzyme engineering studies. We first investigated chalcone synthase (CHS), the entry point of the flavonoid pathway, and its close relative stilbene synthase (STS). Work soon followed on the O-methyl transferases (OMTs) involved in modifications of chalcone, isoflavonoids and metabolic precursors of lignin. More recently, our groups and others have extended the range of phenylpropanoid pathway structural investigations to include the upstream enzymes responsible for the initial recruitment of phenylalanine and tyrosine, as well as a number of reductases, acyltransferases and ancillary tailoring enzymes of phenylpropanoid-derived metabolites. These structure-function studies collectively provide a comprehensive view of an important aspect of phenylpropanoid metabolism. More specifically, these atomic resolution insights into the architecture and mechanistic underpinnings of phenylpropanoid metabolizing enzymes contribute to our understanding of the emergence and on-going evolution of specialized phenylpropanoid products, and underscore the molecular basis of metabolic biodiversity at the chemical level. Finally, the detailed knowledge of the structure, function and evolution of these enzymes of specialized metabolism provide a set of experimental templates for the enzyme and metabolic engineering of production platforms for diverse novel compounds with desirable dietary and medicinal properties.

Bleaching of soda pulp of fibres of Musa textilis nee (abaca) with peracetic acid.

In this work, we studied the influence of operational variables in the bleaching of soda pulp of Musa textilis nee (abaca) [viz. temperature (55-85 degrees C), bleaching time (30-150 min) and peracetic acid concentration oven dry pulp (0.5-4.5%)] on the kappa number and viscosity of the bleached pulp, as well as on the breaking length, burst index and brightness of paper sheets made from it. For this purpose, we used a central composite factorial design in order to identify the optimum operating conditions. In this way equations relating the dependent variables to the operational variables of the bleaching process were derived. These equations reproduce the dependent variables with errors less than 12% for all, except the viscosity which was predicted with errors less than 18%. Obtaining bleached pulp with the highest possible viscosity (1519 ml/g), and paper sheets with the maximum possible breaking length (6547 m) and burst index (5.00 kN/g), entails using a temperature of 55 degrees C, a peracetic acid concentration of 4.5% and a bleaching time of 150 min. This provides a brightness of 79.90%, which is only 6.53% lower than the maximum possible value (85.48%).

Manejo de la depresión en fase de remisión en Atención Primaria / Management of depression in remissionphase in Primary Care

OBJETIVO. La depresión es el problema de salud mental más frecuente en Atención Primaria (AP). Su manejo terapéutico puede ser diferente en AP al del ámbito especialista, sobre todo durante la fase de estabilización. El objetivo del presente estudio fue conocer la actitud del médico de AP frente al paciente depresivo en remisión y evaluar si la resolución total del cuadro es un objetivo factible en este ámbito. MÉTODOS. 1.a parte: encuesta a médicos de AP sobre su experiencia en el manejo de la depresión en remisión. 2.a parte: estudio transversal, multicéntrico, donde se recogieron datos clínicos de pacientes con depresión en fase de remisión, según criterio clínico. Cada médico debía recoger datos de 5 pacientes, de forma consecutiva, durante 3 meses. Se evaluó la proporción de pacientes con remisión total según escala HDRS, versión reducida de 6 ítems (remisión total: HDRS-6 ≤5), y los factores relacionados con un peor estado clínico mediante un test de regresión lineal. RESULTADOS. 1.a parte: la mayoría de entrevistados coincidía en la necesidad de un tratamiento prolongado (entre 6 meses y 1 año), con el mismo fármaco y dosis a las que respondió el paciente. La baja adherencia fue referida como el principal problema de esta etapa. 2.a parte: el 37% de pacientes reunían criterios de remisión total. Edad, ocupación, tiempo de evolución, comorbilidad psiquiátrica, antecedentes depresivos o satisfacción al tratamiento se relacionaron con un peor estado clínico. CONCLUSIONES. El manejo clínico de la depresión en fase de remisión en AP coincide con las guías terapéuticas vigentes. La proporción de pacientes con remisión total fue similar a la observada en otros ámbitos clínicos

OBJECTIVE. Depression is the most common problem of psychiatric disorders encountered in Primary Care. Its management can differ from the psychiatrist's management, mainly during the remission phase. This study aimed to explore attitudes on management of depressed patients in remission in Primary Care and evaluate whether full remission is a feasible objective in this setting. METHODS. Part 1: semi-structured interview to Primary Care physicians on the management of depression in remission. Part 2: cross-sectional, multicenter study. Clinical data from depressed patients in remission (according to physician's criterion) were collected. Each physician should collect data on 5 consecutive patients seen over 3 months. The percentage of patients with full remission was evaluated by the Hamilton Depression Rating scale (6 items-short version) (full remission: HDRS-6 ≤5). Factors related with a worst clinical condition were evaluated by a lineal regression test. RESULTS. Part 1: most of the physicians agreed with the requirement to continue full-dose maintenance therapy for 6 to 12 months. Low adherence to treatment was reported as the main problem in this period. Part 2: 37% of patients met the full-remission criteria. Age, occupation, episode duration, psychiatric comorbidity, psychiatric antecedents, and treatment satisfaction were related with worst clinical condition. CONCLUSIONS. The long-term management of depression in Primary Care agrees with the current treatment guidelines. The percentage of patients fulfilling remission criteria was similar to the reports in other clinical settings

Crystal structure of HIV-1 protease in situ product complex and observation of a low-barrier hydrogen bond between catalytic aspartates.

HIV-1 protease is an effective target for designing drugs against AIDS, and structural information about the true transition state and the correct mechanism can provide important inputs. We present here the three-dimensional structure of a bi-product complex between HIV-1 protease and the two cleavage product peptides AETF and YVDGAA. The structure, refined against synchrotron data to 1.65 A resolution, shows the occurrence of the cleavage reaction in the crystal, with the product peptides still held in the enzyme active site. The separation between the scissile carbon and nitrogen atoms is 2.67 A, which is shorter than a normal van der Waal separation, but it is much longer than a peptide bond length. The substrate is thus in a stage just past the G'Z intermediate described in Northrop's mechanism [Northrop DB (2001) Acc Chem Res 34:790-797]. Because the products are generated in situ, the structure, by extrapolation, can give insight into the mechanism of the cleavage reaction. Both oxygens of the generated carboxyl group form hydrogen bonds with atoms at the catalytic center: one to the OD2 atom of a catalytic aspartate and the other to the scissile nitrogen atom. The latter hydrogen bond may have mediated protonation of scissile nitrogen, triggering peptide bond cleavage. The inner oxygen atoms of the catalytic aspartates in the complex are 2.30 A apart, indicating a low-barrier hydrogen bond between them at this stage of the reaction, an observation not included in Northrop's proposal. This structure forms a template for designing mechanism-based inhibitors.

Optimization of pulping conditions of abaca. An alternative raw material for producing cellulose pulp.

The influence of temperature (150-170 degrees C), pulping time (15-45 min) and soda concentration (5-10%) in the pulping of abaca on the yield, kappa, viscosity, breaking length, stretch and tear index of pulp and paper sheets, was studied. Using a factorial design to identify the optimum operating conditions, equations relating the dependent variables to the operational variables of the pulping process were derived that reproduced the former with errors lower than 25%. Using a high temperature, and a medium time and soda concentration, led to pulp that was difficult to bleach (kappa 28.34) but provided acceptable strength-related properties (breaking length 4728 m; stretch 4.76%; tear index 18.25 mN m2/g), with good yield (77.33%) and potential savings on capital equipment costs. Obtaining pulp amenable to bleaching would entail using more drastic conditions than those employed in this work.

X-ray-induced debromination of nucleic acids at the Br K absorption edge and implications for MAD phasing.

Multi-wavelength anomalous dispersion (MAD) using brominated derivatives is considered a common and convenient technique for solving chemically synthesized nucleic acid structures. Here, it is shown that a relatively moderate X-ray dose (of the order of 5 x 10(15) photons mm(-2)) can induce sufficient debromination to prevent structure determination. The decrease in bromine occupancy with radiation dose can be accounted for by a simple exponential, with an estimated rate constant at the absorption-peak wavelength, 7.4 (0.8) MGy, that is not significantly different from its value at the absorption-edge wavelength, 9.2 (2.6) MGy (the given e.s.d.s assess the relative closeness of the two values, not their absolute accuracy, which is probably worse). Chemically, these results (and others) are consistent with bromine cleavage resulting from direct photodissociation and/or from the action of free electrons, rather than from the action of hydroxyl radicals originating from water dissociation. The free bromine species (Br(-)) diffuse too quickly, even in amorphous ice around 100 K, to allow the determination of a diffusion coefficient. From a practical point of view, it is suggested that a single data collection with a crystal consisting of iodinated instead of brominated derivatives could provide both anomalous scattering and SIR phase information by the progressive cleavage of iodine.

FIP: a highly automated beamline for multiwavelength anomalous diffraction experiments.

FIP is a French Collaborating Research Group (CRG) beamline at the European Synchrotron Radiation Facility (ESRF) dedicated exclusively to crystallography of biological macromolecules, with a special emphasis on multiwavelength anomalous diffraction data collection in the 0.7-1.81 A wavelength range. The optics, consisting of long cylindrical grazing-angle mirrors associated with a cryocooled double-crystal monochromator, delivers an optimal beam in the corresponding energy range. The high level of automation, which includes automated crystal centring, automated data-collection management and data processing, makes the use of this beamline very easy. This is illustrated by the large number of challenging structures that have been solved since 1999.

Automated data processing on beamline FIP (BM30A) at ESRF.

Automation of protein crystallography synchrotron beamlines is becoming necessary to face challenging structural genomics projects. In this context, a program has been developed that processes diffraction frames using popular software but analyzes statistics and makes choices the way crystallographers usually do. This program includes the classical peak search, indexing, integration, scaling and anomalous signal analysis. The result, comparable with that obtained by standard users, is rapidly available, providing the required information for a more efficient use of the beam time.

Influence of the operating conditions in the acetone pulping of wheat straw on the properties of the resulting paper sheets.

A central composite factor design was used to examine the influence of independent variables in the acetone pulping of wheat straw (processing temperature, time, and acetone concentration) on the yield of the resulting pulp, and on various physical properties of paper sheets (breaking length, stretch, burst index, tear index and brightness) obtained from it. Equations that related each dependent variable to the different independent variables were obtained, and these reproduced the experimental results for the yield, breaking, stretch, burst index and brightness obtained at temperatures, times, and acetone concentratons over the ranges 140-180 degrees C, 60-120 min and 40-80%, respectively, with errors less than 20%. Obtaining the optimum breakinig length, stretch, burst index and tear index for the paper sheets (3,456 m, 1.42%, 1.36 kN/g and 3.86 mNm2/g, respectively) entails using a high temperature; the processing time and acetone concentration only influence stretch, optimization of which requires using a short time and a low concentration. The optimum brightness (30.44%) is achieved with a low temperature, a short time and a medium acetone concentration. In order to minimize losses of solvent during its recovery and recycling while ensuring acceptable levels of the properties of the paper sheets, a high temperature, a low acetone concentration and a short time can be used; the brightness level thus obtained is only 10% lower than the optimum value.

Structure and mechanism of chalcone synthase-like polyketide synthases.

Polyketide synthases (PKS) produce an array of natural products with different biological activities and pharmacological properties by varying the starter and extender molecules that form the final polyketide. Recent studies of the simplest PKS, the chalcone synthase (CHS)-like enzymes involved in the biosynthesis of flavonoids, anthocyanin pigments, and antimicrobial phytoalexins, have yielded insight on the molecular basis of this biosynthetic versatility. Understanding the structure-function relationship in these PKS provides a foundation for manipulating polyketide formation and suggests strategies for further increasing the scope of polyketide biosynthetic diversity.

Four crystal structures of the 60 kDa flavoprotein monomer of the sulfite reductase indicate a disordered flavodoxin-like module.

Escherichia coli NADPH-sulfite reductase (SiR) is a 780 kDa multimeric hemoflavoprotein composed of eight alpha-subunits (SiR-FP) and four beta-subunits (SiR-HP) that catalyses the six electron reduction of sulfite to sulfide. Each beta-subunit contains a Fe4S4 cluster and a siroheme, and each alpha-subunit binds one FAD and one FMN as prosthetic groups. The FAD gets electrons from NADPH, and the FMN transfers the electrons to the metal centers of the beta-subunit for sulfite reduction. We report here the 1.94 A X-ray structure of SiR-FP60, a recombinant monomeric fragment of SiR-FP that binds both FAD and FMN and retains the catalytic properties of the native protein. The structure can be divided into three domains. The carboxy-terminal part of the enzyme is composed of an antiparallel beta-barrel which binds the FAD, and a variant of the classical pyridine dinucleotide binding fold which binds NADPH. These two domains form the canonic FNR-like module, typical of the ferredoxin NADP+ reductase family. By analogy with the structure of the cytochrome P450 reductase, the third domain, composed of seven alpha-helices, is supposed to connect the FNR-like module to the N-terminal flavodoxine-like module. In four different crystal forms, the FMN-binding module is absent from electron density maps, although mass spectroscopy, amino acid sequencing and activity experiments carried out on dissolved crystals indicate that a functional module is present in the protein. Our results clearly indicate that the interaction between the FNR-like and the FMN-like modules displays lower affinity than in the case of cytochrome P450 reductase. The flexibility of the FMN-binding domain may be related, as observed in the case of cytochrome bc1, to a domain reorganisation in the course of electron transfer. Thus, a movement of the FMN-binding domain relative to the rest of the enzyme may be a requirement for its optimal positioning relative to both the FNR-like module and the beta-subunit.

Dissection of malonyl-coenzyme A decarboxylation from polyketide formation in the reaction mechanism of a plant polyketide synthase.

Chalcone synthase (CHS) catalyzes formation of the phenylpropanoid chalcone from one p-coumaroyl-CoA and three malonyl-coenzyme A (CoA) thioesters. The three-dimensional structure of CHS [Ferrer, J.-L., Jez, J. M., Bowman, M. E., Dixon, R. A., and Noel, J. P. (1999) Nat. Struct. Biol. 6, 775-784] suggests that four residues (Cys164, Phe215, His303, and Asn336) participate in the multiple decarboxylation and condensation reactions catalyzed by this enzyme. Here, we functionally characterize 16 point mutants of these residues for chalcone production, malonyl-CoA decarboxylation, and the ability to bind CoA and acetyl-CoA. Our results confirm Cys164's role as the active-site nucleophile in polyketide formation and elucidate the importance of His303 and Asn336 in the malonyl-CoA decarboxylation reaction. We suggest that Phe215 may help orient substrates at the active site during elongation of the polyketide intermediate. To better understand the structure-function relationships in some of these mutants, we also determined the crystal structures of the CHS C164A, H303Q, and N336A mutants refined to 1.69, 2.0, and 2.15 A resolution, respectively. The structure of the C164A mutant reveals that the proposed oxyanion hole formed by His303 and Asn336 remains undisturbed, allowing this mutant to catalyze malonyl-CoA decarboxylation without chalcone formation. The structures of the H303Q and N336A mutants support the importance of His303 and Asn336 in polarizing the thioester carbonyl of malonyl-CoA during the decarboxylation reaction. In addition, both of these residues may also participate in stabilizing the tetrahedral transition state during polyketide elongation. Conservation of the catalytic functions of the active-site residues may occur across a wide variety of condensing enzymes, including other polyketide and fatty acid synthases.