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The (S)-norcoclaurine synthase from Thalictrum flavum(TfNCS) stereoselectively catalyzes the Pictet–Spengler reaction between dopamine and 4-hydroxyphenylacetaldehyde to give (S)-norcoclaurine. TfNCS can catalyze the Pictet–Spengler reaction with various aldehydes and ketones, leading to diverse tetrahydroisoquinolines. This substrate promiscuity positions TfNCS as a highly promising enzyme for synthesizing fine chemicals. Understanding carbonyl-containing substrates’ structural and electronic signatures that influence TfNCS activity can help expand its applications in the synthesis of different compounds and aid in protein optimization strategies. In this study, we investigated the influence of the molecular properties of aldehydes and ketones on their reactivity in the TfNCS-catalyzed Pictet–Spengler reaction. Initially, we compiled a library of reactive and unreactive compounds from previous publications. We also performed enzymatic assays using nuclear magnetic resonance to identify some reactive and unreactive carbonyl compounds, which were then included in the library. Subsequently, we employed QSAR and DFT calculations to establish correlations between substrate-candidate structures and reactivity. Our findings highlight correlations of structural and stereoelectronic features, including the electrophilicity of the carbonyl group, to the reactivity of aldehydes and ketones toward the TfNCS-catalyzed Pictet–Spengler reaction. Interestingly, experimental data of seven compounds out of fifty-three did not correlate with the electrophilicity of the carbonyl group. For these seven compounds, we identified unfavorable interactions between them and the TfNCS. Our results demonstrate the applications of in silicotechniques in understanding enzyme promiscuity and specificity, with a particular emphasis on machine learning methodologies, DFT electronic structure calculations, and molecular dynamic (MD) simulations.

This work reports the catalysis of bare silica materials (MCM-41, SBA-15, and silica gel) for the complete hydrolyses of bis(2,4-nitrophenyl)phosphate (BDNPP) and bis(4-nitrophenyl)phosphate (BNPP), which share the same phosphate diester chemical core with important biological molecules, DNA and RNA, and toxic substances like EA2192 and anatoxin-a(s). Catalytic effects above 7000-fold were observed for the hydrolysis of BNPP, which were consistent with a concerted SN2(P) mechanism under the action of siloxide and silanol groups that provide water activation by general base catalysis and leaving-group departure stabilization by general acid catalysis, respectively. We also observed that product formation follows a sigmoidal behavior as a result of catalytic effect enhancement following an induction period. This effect is discussed considering cooperative effects caused by silica hydration, which is unchanged upon material reuse affording strong catalytic effects for phosphate diester hydrolyses and no induction periods.

•The importance of dynamics in proteins complexes formation is discussed.•Specific and nonspecific interactions in recognition.•Conformational ensemble in the conformational selection mechanisms.