Aromatic Amino Acid Residues Research Articles

Construction of functional soybean peptide–cyclodextrin carboxylate nanoparticles and their interaction with porcine pancreatic α-amylase

In this study, soybean peptide-succinic acid-modified cyclodextrin (SPT-SACD) nanoparticles (NPs) were successfully fabricated by combining SPT and SACD using an antisolvent precipitation approach. The effects of the average molecular weight of SPT and the SPT/SACD mass ratio on the structure and properties of the SACD-SPT NPs were investigated. Under optimal conditions, the SPT/SACD mass ratio was 2:1, and the SPT average molecular weight was 300 Da. SPT-SACD NPs were prepared under these conditions were spherical and had good uniformity. The particle sizes by DLS of SPT1 (300 Da) /SACD and SPT2 (500 Da) /SACD were in the range of 250–400 nm. The interaction between α-amylase and SPT-SACD NPs was investigated using ultraviolet visible (UV–Vis) absorption, fluorescence, and circular dichroism (CD) spectroscopy. The results of the fluorescence spectra and CD spectroscopy suggested that the presence of SPT-SACD NPs changed the microenvironment of the aromatic amino acid residues, which leads to the change of enzyme protein structure. The SPT-SACD NPs statically quenched the intrinsic fluorescence of the α-amylase by forming a complex with the enzyme. Moreover, the SPT-SACD NPs significantly improved the inhibitory effect of EGCG on α-amylase. The semi-inhibitory concentration (IC50) decreased from 0.324 to 0.248 mg/mL. This study provides an improved understanding of the interaction mechanism between polypeptide-cyclodextrin complexes and digestive enzymes, which may facilitate the design of functional foods.

Effect of Laser-Induced Optical Breakdown on the Structure of Bsa Molecules in Aqueous Solutions: An Optical Study.

The influence of laser radiation of a typical surgical laser on the physicochemical properties of the Bovine Serum Albumin (BSA) protein was studied. It was established that the physicochemical characteristics of optical breakdown weakly depend on the concentration of protein molecules. At the same time, the patterns observed for an aqueous solution of BSA irradiated with a laser for different time periods were extremely similar to the classical ones. It was established that after exposure to laser radiation, the optical density of protein solutions increases. At the same time, the intensity of BSA fluorescence due to aromatic amino acid residues decreases insignificantly after exposure to laser radiation. In this case, the position of the excitation and emission maximum does not change, and the shape of the fluorescence spot on 3D maps also does not change significantly. On the Raman spectrum after exposure to laser radiation, a significant decrease in 1570 cm−1 was observed, which indicates the degradation of α-helices and, as a result, partial denaturation of BSA molecules. Partial denaturation did not significantly change the total area of protein molecules, since the refractive index of solutions did not change significantly. However, in BSA solutions, after exposure to laser radiation, the viscosity increased, and the pseudoplasticity of aqueous solutions decreased. In this case, there was no massive damage to the polypeptide chain; on the contrary, when exposed to optical breakdown, intense aggregation was observed, while aggregates with a size of 400 nm or more appeared in the solution. Thus, under the action of optical breakdown induced by laser radiation in a BSA solution, the processes of partial denaturation and aggregation prevail, aromatic amino acid residues are damaged to a lesser extent, and fragmentation of protein molecules is not observed.

The Spatial Distribution Patterns, Physicochemical Properties, and Structural Characterization of Proteins in Oysters (Crassostrea hongkongensis).

Protein content, a vital component determining the nutritional quality of oysters, is unevenly distributed in different parts of oyster. In this study, the spatial distribution (visceral mass, mantle, gill, and adductor) patterns and structural characteristics of proteins, including water–soluble proteins (WSP), salt–soluble proteins (SSP), acid–soluble proteins (ASP) and alkali–soluble proteins (ALSP) of oysters (Crassostrea hongkongensis) were investigated with the amino acid analyzer, circular dichroism spectroscopy (CD), fourier transform infrared spectroscopy (FTIR), and fluorescence spectroscopy. The results showed that oyster proteins were mainly distributed in the visceral mass and mantle. The protein composition was WSP, SSP, ALSP, and ASP in descending order, which conformed to the ideal amino acid pattern. Variations in secondary structure, molecular weight distribution, and thermal denaturation temperatures of the oyster proteins were observed. SSP had wider bands (16–270 kDa) than those of ASP (30–37 kDa) and ALSP (66–270 kDa). Among the four proteins, the SSP of the mantle showed the highest thermal stability (87.4 °C), while ALSP of the adductor muscle had the lowest the lowest the peak denaturation temperature (Tm) (53.8 °C). The proportions of secondary structures in oyster proteins were different, with a higher proportion of solid protein β–folds, and the exposure of aromatic amino acid residues and disulfide bonds and the microenvironment in which they were located were also different.

Structure-activity relationships and the underlying mechanism of α-amylase inhibition by hyperoside and quercetin: Multi-spectroscopy and molecular docking analyses

Inhibiting the activity of α-amylase has been considered an effective strategy to manage hyperglycemia. Hyperoside and quercetin are the main natural flavonoids in various plants, and the inhibition mechanism on α-amylase remains unclear. In this study, the structure–activity relationships between hyperoside/quercetin and α-amylase were evaluated by enzyme kinetic analysis, multi-spectroscopic techniques, and molecular docking analysis. Results showed that hyperoside and quercetin exhibited significant α-amylase inhibitory activities with IC50 values of 0.491 and 0.325 mg/mL, respectively. The α-amylase activity decreased in the presence of hyperoside and quercetin in a competitive and noncompetitive manner, respectively. UV–vis spectra suggested that the aromatic amino acid residues (Trp and Tyr) microenvironment of α-amylase changed in the presence of these two flavonoids. FTIR and CD spectra showed the vibrations of the amide bands and the secondary structure content changes. The fluorescence quenching mechanism of α-amylase by hyperoside and quercetin belonged to the static quenching type. Finally, molecular docking intuitively showed that hyperoside/quercetin formed hydrogen bonds with the key active site residues (Asp197, Glu233, and Asp300) in α-amylase. MD simulation indicated hyperoside/quercetin-α-amylase docked complexes had good stability. Taken together, this research provides new sights to developing potent drugs or functional foods with hyperoside and quercetin, offering new avenues for hyperglycemia treatment.

Total Internal Reflection Tip-Enhanced Raman Spectroscopy of Tau Fibrils.

Total internal reflection tip-enhanced Raman spectroscopy (TIR-TERS) has recently emerged as a promising technique for noninvasive nanoscale chemical characterization of biomolecules. We demonstrate that the TERS enhancement achieved in this experimental configuration is nearly 30 times higher than that in linear polarization and 8 times higher than that in radial polarization using traditional bottom-illumination geometry. TIR-TERS is applied to the study of Tau amyloid fibrils formed with the human full-length Tau protein mixed with heparin. This technique reveals the possibility to perform TERS imaging with 1-4 nm axial and 5-10 nm lateral spatial optical resolution. In these Tau/heparin fibrils, spectral signatures assigned to aromatic amino acid residues (phenylalanine, histidine, and tyrosine) and nonaromatic ones (e.g., cysteine, lysine, arginine, asparagine, and glutamine) are distinctly observed. Amide I and amide III bands can also be detected. In a fibril portion, it is shown that antiparallel β-sheets and fibril core β-sheets are abundant and are often localized in amino acid-rich regions where parallel β-sheets and random coils are present in lower proportions. This first TIR-TERS study on a nonresonant biological sample paves the way for future nanoscale chemical and structural characterization of biomolecules using this performant and original technique.

Distinctive features of the respiratory syncytial virus priming loop compared to other non-segmented negative strand RNA viruses.

De novo initiation by viral RNA-dependent RNA polymerases often requires a polymerase priming residue, located within a priming loop, to stabilize the initiating NTPs. Polymerase structures from three different non-segmented negative strand RNA virus (nsNSV) families revealed putative priming loops in different conformations, and an aromatic priming residue has been identified in the rhabdovirus polymerase. In a previous study of the respiratory syncytial virus (RSV) polymerase, we found that Tyr1276, the L protein aromatic amino acid residue that most closely aligns with the rhabdovirus priming residue, is not required for RNA synthesis but two nearby residues, Pro1261 and Trp1262, were required. In this study, we examined the roles of Pro1261 and Trp1262 in RNA synthesis initiation. Biochemical studies showed that substitution of Pro1261 inhibited RNA synthesis initiation without inhibiting back-priming, indicating a defect in initiation. Biochemical and minigenome experiments showed that the initiation defect incurred by a P1261A substitution could be rescued by factors that would be expected to increase the stability of the initiation complex, specifically increased NTP concentration, manganese, and a more efficient promoter sequence. These findings indicate that Pro1261 of the RSV L protein plays a role in initiation, most likely in stabilizing the initiation complex. However, we found that substitution of the corresponding proline residue in a filovirus polymerase had no effect on RNA synthesis initiation or elongation. These results indicate that despite similarities between the nsNSV polymerases, there are differences in the features required for RNA synthesis initiation.

Effects of different cooking methods on peanut allergenicity

Peanut allergy is a type of serious food allergy worldwide. In this study, peanuts were treated with two different cooking methods. The effect of cooking on digestion stability was evaluated by simulated gastrointestinal digestions. The effects of cooking on the allergenicity were assessed by immunoblotting, passive cutaneous anaphylaxis (PCA) mice model and rat basophilic leukemia (RBL)-2H3 cell model. Results showed that raw peanuts were more stable to digestion than boiled and fried peanuts. SDS-PAGE showed that the quantities of allergens Ara h 1 (64 kDa), Ara h 2 (20 kDa), and Ara h 3 (38 kDa) in the boiled and fried peanuts were all reduced. Meanwhile, immunoblotting revealed the IgE binding capacities of boiled and fried peanuts were decreased significantly, especially the fried peanuts. In the PCA mice model, the vascular permeability of allergic mice, as well as the release of histamine, were both alleviated in mice gavaged with boiled and fried peanuts compared with raw peanuts. Lastly, the release of β-hexosaminidase of RBL-2H3 cells treated with boiled and fried peanuts was significantly decreased compared with the raw peanuts. In addition, the analyses of circular dichroism (CD) spectra showed that the α-helical content of peanut proteins was decreased after boiling and frying, indicating the structure stability of peanut allergens decreased. The analyses of ultraviolet (UV) spectra showed that aromatic amino acid residues were exposed, causing the change of the tertiary structure of peanut allergens after boiling and frying. In conclusion, the allergenicity of peanuts was significantly reduced after boiling and frying, especially frying.

Physicochemical characterisation of ACE‐inhibitory and antioxidant peptides from Alcalase® whey protein hydrolysates using fractionation strategies

Antioxidant activity (AOA) and ACE‐inhibitory activity (AHA) were obtained by enzymatic hydrolysis of bovine whey using food‐grade Alcalase®. The aim of this report was to characterise relevant physicochemical properties of the bioactive peptides and evaluate them with respect to the degree of hydrolysis (DH) using separation techniques. The results suggest that low molecular weight peptides containing high concentrations of cationic and aromatic amino acid residues were responsible for AHA, whereas those related to AOA would possess anionic and both nonpolar aliphatic and aromatic amino acid residues within their sequence. Furthermore, an extensive DH negatively influenced radical scavenging AOA but positively affected angiotensin‐converting enzyme (ACE) inhibition.

Hybrid-Integrated Biosensor for Express Determination of Protein Markers of Diseases based on Molecular Recognition and Direct Fluorimetric Detection

Ways of creating new generation biosensors for multiparametric express diagnostics based on molecular recognition and direct fluorimetric registration of a peptide aptamer — protein marker complex were considered. The biosensor platform comprises a microfluidic channel for delivery sample solutions, coupled with flow-through zones containing covalently attached arrays of peptide probes — aptamers. An outer glass window of the biochip assembly contains a layer of luminophore ZnS:Cu, bound on it via an acrylic lacquer and intended for the re-emitting native fluorescence of bound proteins into the longer wavelength range, more efficient in registering signals with CMOS sensors. The aptamers were designed using "Protein 3D" program for analysis of spatial complementarity of protein structures. The peptide, complementary to Troponin T, was modified by replacement of aromatic amino acid residue while maintaining the spatial configuration. The complementarity of peptide and Troponin T was confirmed using a capillary electrophoresis-on-a-chip. Biosensors are manufactured using thick-film technology and photolithography. The fluorescence of marker proteins was excited using UV-LED with a radiation wavelength of 275 nm. The limit of detection achieved for Troponin T was 6 ng/ml.

Clustering of Aromatic Amino Acid Residues around Methionine in Proteins.

Short-range, non-covalent interactions between amino acid residues determine protein structures and contribute to protein functions in diverse ways. The interactions of the thioether of methionine with the aromatic rings of tyrosine, tryptophan, and/or phenylalanine has long been discussed and such interactions are favorable on the order of 1–3 kcal mol−1. Here, we carry out a new bioinformatics survey of known protein structures where we assay the propensity of three aromatic residues to localize around the [-CH2-S-CH3] of methionine. We term these groups “3-bridge clusters”. A dataset consisting of 33,819 proteins with less than 90% sequence identity was analyzed and such clusters were found in 4093 structures (or 12% of the non-redundant dataset). All sub-classes of enzymes were represented. A 3D coordinate analysis shows that most aromatic groups localize near the CH2 and CH3 of methionine. Quantum chemical calculations support that the 3-bridge clusters involve a network of interactions that involve the Met-S, Met-CH2, Met-CH3, and the π systems of nearby aromatic amino acid residues. Selected examples of proposed functions of 3-bridge clusters are discussed.

Analysis of the isomerization of diketopiperazine consisting of proline and aromatic amino acid residues using nuclear magnetic resonance.

Under different concentrations of the base potassium deuteroxide KOD, the progress of reactions, such as enolization, D-substitution, isomerization, and conformational changes of diketopiperazine cyclo(L-Pro-L-Xxx) and cyclo(D-Pro-L-Xxx) (Xxx=Phe, Tyr) in D2O solution, was investigated by 1H nuclear magnetic resonance (NMR). Cyclo(L-Pro-L-Xxx) is mostly isomerized to cyclo(D-Pro-L-Xxx) in D2O solution, whereas cyclo(D-Pro-L-Xxx) is only slightly isomerized to cyclo(L-Pro-L-Xxx) even under stronger basic conditions. After adding a deuterated organic solvent (CD3COCD3, CD3SOCD3 or CD3OD) to a D2O solution of cyclo(L-Pro-L-Xxx), cyclo(D-Pro-L-Xxx), or increasing the temperature of the D2O solution, CH-π interaction between H9 and the benzene ring of cyclo (D-Pro-L-Xxx) was stronger than that between H8α and the benzene ring of cyclo(L-Pro-L-Xxx).

Nano‐Assemblies of a Synthetic Peptide: Illuminating Aggregation Potential, Amyloidogenicity and Cytotoxicity

AbstractProtein misfolding and subsequent generation of amyloid fibrils are associated with a wide range of diseases e. g. Alzheimer's, Huntington's, Parkinson's, type II diabetes, cystic fibrosis, and many others. Aromatic amino acid residues of polypeptide/protein are supposed to play a critical role in formation and stabilization of amyloid fibrils. Herein, we report a synthetic peptide, ALYLV (peptide 1), corresponding to B14‐18 of insulin, that forms nanofibrillar aggregates upon self‐association in aqueous medium. Staining with congo red (CR) and thioflavin t (ThT) establish that these organized nano‐assemblies are amyloidogenic in nature. Interestingly, the structural modification of peptide 1 replacing the centrally located tyrosine residue with valine, ALVLV (peptide 2), significantly reduces the assembly propensity as well as the amyloidogenicity of peptide 2 compared to peptide 1. MTT (3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide) cell survival assay reveals the cytotoxicity of the aggregated peptides on HeLa cells upon 48 h of exposure. Overall, the study explores the aggregation potential, amyloidogenicity, and cytotoxicity of a hotspot of insulin and pinpoints the role of aromatic amino acid residue in amyloidosis. This outcome also illuminates the peptide nano‐assemblies for various biotechnological applications.

Unraveling the Compositional and Molecular Features Involved in Lysozyme-Benzothiazole Derivative Interactions.

In this work we present a computational analysis together with experimental studies, focusing on the interaction between a benzothiazole (BTS) and lysozyme. Results obtained from isothermal titration calorimetry, UV-vis, and fluorescence were contrasted and complemented with molecular docking and machine learning techniques. The free energy values obtained both experimentally and theoretically showed excellent similarity. Calorimetry, UV-vis, and 3D/2D-lig-plot analysis revealed that the most relevant interactions between BTS and lysozyme are based on a predominance of aromatic, hydrophobic Van der Waals interactions, mainly aromatic edge-to-face (T-shaped) π-π stacking interactions between the benzene ring belonging to the 2-(methylthio)-benzothiazole moiety of BTS and the aromatic amino acid residue TRP108 of the lysozyme receptor. Next, conventional hydrogen bonding interactions contribute to the stability of the BTS-lysozyme coupling complex. In addition, mechanistic approaches performed using elastic network models revealed that the BTS ligand theoretically induces propagation of allosteric signals, suggesting non-physiological conformational flexing in large blocks of lysozyme affecting α-helices. Likewise, the BTS ligand interacts directly with allosteric residues, inducing perturbations in the conformational dynamics expressed as a moderate conformational softening in the α-helices H1, H2, and their corresponding β-loop in the lysozyme receptor, in contrast to the unbound state of lysozyme.

Estimation of the selectivity of sorbents for IgG binding based on tripeptide ligands

Biospecific sorbents for the removal of IgG and subclasses from biological fluids based on oligopeptides, containing aromatic amino acid residues, were created. The selectivity properties of specific sorbents for IgM, IgE, and plasma proteins were evaluated. It was found that the created sorbents exhibit the low activity to the total plasma protein, albumin, IgM, IgE and are highly specific for IgG.

An “up” oriented methionine-aromatic structural motif in SUMO is critical for its stability and activity

Protein structural bioinformatic analyses suggest preferential associations between methionine and aromatic amino acid residues in proteins. Ab initio energy calculations highlight a conformation-dependent stabilizing interaction between the interacting sulfur-aromatic molecular pair. However, the relevance of buried methionine-aromatic motifs to protein folding and function is relatively unexplored. The Small Ubiquitin-Like Modifier (SUMO) is a β-grasp fold protein and a common posttranslational modifier that affects diverse cellular processes, including transcriptional regulation, chromatin remodeling, metabolic regulation, mitosis, and meiosis. SUMO is a member of the Ubiquitin-Like (UBL) protein family. Herein, we report that a highly conserved and buried methionine-phenylalanine motif is a unique signature of SUMO proteins but absent in other homologous UBL proteins. We also detect that a specific “up” conformation between the methionine-phenylalanine pair of interacting residues in SUMO is critical to its β-grasp fold. The noncovalent interactions of SUMO with its ligands are dependent on the methionine–phenylalanine pair. MD simulations, NMR, and biophysical and biochemical studies suggest that perturbation of the methionine-aromatic motif disrupts native contacts, modulates noncovalent interactions, and attenuates SUMOylation activity. Our results highlight the importance of conserved orientations of Met-aromatic structural motifs inside a protein core for its structure and function.

Toward Development of a Label-Free Detection Technique for Microfluidic Fluorometric Peptide-Based Biosensor Systems.

The problems of chronic or noncommunicable diseases (NCD) that now kill around 40 million people each year require multiparametric combinatorial diagnostics for the selection of effective treatment tactics. This could be implemented using the biosensor principle based on peptide aptamers for spatial recognition of corresponding protein markers of diseases in biological fluids. In this paper, a low-cost label-free principle of biomarker detection using a biosensor system based on fluorometric registration of the target proteins bound to peptide aptamers was investigated. The main detection principle considered includes the re-emission of the natural fluorescence of selectively bound protein markers into a longer-wavelength radiation easily detectable by common charge-coupled devices (CCD) using a specific luminophore. Implementation of this type of detection system demands the reduction of all types of stray light and background fluorescence of construction materials and aptamers. The latter was achieved by careful selection of materials and design of peptide aptamers with substituted aromatic amino acid residues and considering troponin T, troponin I, and bovine serum albumin as an example. The peptide aptamers for troponin T were designed in silico using the «Protein 3D» (SPB ETU, St. Petersburg, Russia) software. The luminophore was selected from the line of ZnS-based solid-state compounds. The test microfluidic system was arranged as a flow through a massive of four working chambers for immobilization of peptide aptamers, coupled with the optical detection system, based on thick film technology. The planar optical setup of the biosensor registration system was arranged as an excitation-emission cascade including 280 nm ultraviolet (UV) light-emitting diode (LED), polypropylene (PP) UV transparent film, proteins layer, glass filter, luminophore layer, and CCD sensor. A laboratory sample has been created.

Quinoa bioactive protein hydrolysate produced by pancreatin enzyme- functional and antioxidant properties

Quinoa protein with great functional properties has been considered a potential source for food fortification or the production of functional foods. In this study, quinoa protein hydrolysate (QPH) was prepared from quinoa protein concentrate (QPC) after hydrolysis by pancreatin (pancreatic) enzyme. The emulsifying, foaming, and antioxidant activities, the emulsion and foam stabilization, as well as amino acid profile and chemical composition were evaluated. The highest degree of hydrolysis (19.17%) was obtained after 180 min. The amino acid profile (by high-performance liquid chromatography: HPLC) showed that glutamic acid and lysine had the highest contents and all essential (limiting) amino acids were recorded. Fourier-transform infrared (FTIR) spectroscopy showed hydroxylic and aromatic amino acid residues in the QPC and it was inferred that different functional groups have originated in QPH that confirms the hydrolyzing action of pancreatin enzyme. QPH had the lowest solubility at the isoelectric pH of 5 (pI~5) and had better emulsifying, foaming, and antioxidant properties compared to QPC. In contrast, QPC had better emulsion/foam stabilizing properties. Overall, this study suggests that pancreatin could be employed for hydrolysis of quinoa protein and its hydrolysate with functional properties could be possibly considered for food fortification or production of functional foods.

Simultaneous quantification of total carbohydrate and protein amounts from aqueous solutions by the sulfuric acid ultraviolet absorption method (SA-UV method)

Based on the sulfuric acid-ultraviolet assay (SA-UV, developed by Albalasmeh et al., 2013), we have further expanded this method for the simultaneous quantification of saccharides (carbohydrates) and proteins by ultraviolet spectrophotometry. The absorbance of saccharides depends on the formation of furfurals by dehydration in the presence of concentrated sulfuric acid, whereas proteins are unaffected and can be quantified by UV active peptide bonds and aromatic amino acid residues. In saccharide/protein mixtures the SA-UV assay offers a good alternative and substitutes the need for two different methods, like the phenol-sulfuric acid (PSA, developed by DuBois et al., 1951) and bicinchoninic acid (BCA, developed by Smith et al., 1985) assays. For the development of this method, we used glucose and BSA as model substrates and performed a method validation in terms of linearity, LOD, LOQ, accuracy, and precision. Simultaneous quantification in glucose/BSA mixtures is possible down to 20 mg/L from 30 μL sample volumes, and even low content mixtures with concentrations down to 2 mg/L can appropriately be quantified from higher volumes by an evaporation technique.

Peptides on a rescue mission against the hazardous cidal chemicals used in Eastern Indian Agriculture: An in-silico approach based on field survey in 2020

Background: The use of poisonous agrochemicals is both necessary and menacing for a farmer. The long-term hazards of chemicals can even be life-threatening to many people. Recently, the adverse effects of such chemicals on the environment and human health received serious attention throughout the world. In an Indian context, the usage of the chemical sprays has costed many lives and influenced vital organs to the greatest extent. This serious situation advocates the search for ways to prevent the inhalation and absorption of these chemicals into the body. The present study was conducted to evaluate different types of chemical sprays, their impact, and explore remedial ways to neutralize their toxic effects. Methods: Alpha-cypermethrin, phorate, fenobucarb, buprofezin, and mancozeb were selected for binding based on the feedback of severity in actions reported by the respondents and statistical analysis of different physical and chemical parameters. Chemical similarity analysis of the 39 cidal chemicals was performed to predict the binding similarity with respect to the above-mentioned 5 seed molecules. These five chemicals were docked with three different proteins 121p GTPase HRas, Torpedo acetylcholinesterase, and trypanosoma phosphoglycerate kinase, selected based on the common target prediction. Results: Common target prediction revealed that these chemicals have an affinity to bind these proteins with a higher binding interaction towards the aromatic amino acid residues and those capable of H-bonding, escorted by a predilection towards acetylcholinesterase. Conclusion: The in-silico findings are the basis to propose that acetylcholinesterase can be considered as a potential binding agent to trap fatal agrochemicals.

Rerouting and Improving Dauc-8-en-11-ol Synthase from Streptomyces venezuelae to a High Yielding Biocatalyst.

The dauc‐8‐en‐11‐ol synthase from Streptomyces venezuelae was investigated for its catalytic activity towards alternative terpene precursors, specifically designed to enable new cyclisation pathways. Exchange of aromatic amino acid residues at the enzyme surface by site‐directed mutagenesis led to a 4‐fold increase of the yield in preparative scale incubations, which likely results from an increased enzyme stability instead of improved enzyme kinetics.