Shyamalava Mazumdar

Metal‐porphyrin‐based biocatalysts are gaining popularity in sustainable chemical synthesis. Heme‐based enzymes like myoglobins, cytochrome P450s are known for their regio‐ and stereo‐selective cyclopropanation reactions in aqueous media. However, their practical application is challenged by complex, multistep synthesis procedures and inherent structural instability. To address these limitations, we developed a stable, enzyme‐mimicking heterogeneous catalyst using porous SBA15 material. The inner core of SBA15 was functionalized with an alkyl chain and protoporphyrin IX iron (III) (hemin), resulting in alkyl‐SBA15‐amideHemin. Characterization through TEM, UV–vis and FT‐IR spectroscopy, CHN analysis etc., confirmed the successful immobilization of hemin within the hydrophobic pores of functionalized SBA15. The catalytic performance of alkyl‐SBA15‐amideHemin was evaluated in the cyclopropanation reaction of styrene with ethyl diazoacetate (EDA), achieving a turnover number (TON) of 144—nearly double that of free hemin (TON 75). The catalyst exhibited high trans diastereoselectivity (∼70%) for 2‐(cyclopropyl benzene) esters in aqueous media, outperforming other known heterogeneous catalysts. Its catalytic activity and selectivity were maintained over four recycling cycles, highlighting its stability and reusability. Steady‐state emission studies, as well as the kinetics of the iron‐carbene complex, provided insights into the catalytic mechanism. These findings suggest that immobilized hemin within a hydrophobic environment serves as a robust and selective catalyst for chemical transformations in aqueous media.

Metal-porphyrin-based biocatalysts are gaining popularity in sustainable chemical synthesis. Heme-based enzymes like myoglobins, cytochrome P450s are known for their regio- and stereo-selective cyclopropanation reactions in aqueous media. However, their practical application is challenged by complex, multistep synthesis procedures and inherent structural instability. To address these limitations, we developed a stable, enzyme-mimicking heterogeneous catalyst using porous SBA-15 material. The inner core of SBA-15 was functionalized with an alkyl chain and protoporphyrin IX iron (III) (hemin), resulting in alkyl-SBA15-amideHemin. Characterization through TEM, UV-Vis, and FT-IR confirmed the successful immobilization of hemin within the hydrophobic pores of functionalized SBA-15. The catalytic performance of alkyl-SBA15-amideHemin was evaluated in the cyclopropanation reaction of styrene with ethyl diazoacetate (EDA), achieving a turnover number (TON) of 144—nearly double that of free hemin (TON 75). The catalyst exhibited high trans diastereoselectivity (~70%) for 2-(cyclopropyl benzene) esters in aqueous media, outperforming other known heterogeneous catalysts. Its catalytic activity and selectivity were maintained over four recycling cycles, highlighting its stability and reusability. Steady-state emission studies of the iron-carbene complex provided insights into the catalytic mechanism. These findings suggest that immobilized hemin within a hydrophobic environment serves as a robust and selective catalyst for chemical transformations in aqueous media.

Rational design and engineering of the active pocket for access and binding of the substrate, and water channel in a thermostable cytochrome P450 enabled efficient catalysis of a longer chain alkane.

The role of the shape of the nanostructure on the antibacterial effects of ZnO nanodisks has been investigated by detailed mass spectrometry-based proteomics along with other spectroscopic and microscopic studies on E. coli. The primary interaction study of the E. coli cells in the presence of ZnO nanodisks showed rigorous cell surface damage disrupting the cell wall/membrane components detected by microscopic and ATR-FTIR studies. Protein profiling of whole-cell extracts in the presence and absence of ZnO nanodisks identified several proteins that are upregulated and downregulated under the stress of the nanodisks. This suggests that the bacterial response to the primary stress leads to a secondary impact of ZnO nanodisk toxicity via regulation of the expression of specific proteins. Results showed that the ZnO nanodisks lead to the over-expression of peptidyl-dipeptidase Dcp, Transketolase-1, etc., which are important to maintaining the osmotic balance in the cell. The abrupt change in osmotic pressure leads to mechanical injury to the membrane, and nutritional starvation conditions, which is revealed from the expression of the key proteins involved in membrane-protein assembly, maintaining membrane integrity, cell division processes, etc. Thus, indicating a deleterious effect of ZnO nanodisk on the protective layer of E. coli. ZnO nanodisks seem to primarily affect the protective membrane layer, inducing cell death via the development of osmotic shock conditions, as one of the possible reasons for cell death. These results unravel a unique behavior of the disk-shaped ZnO nanostructure in executing lethality in E. coli, which has not been reported for other known shapes or morphologies of ZnO nanoforms.

Thermostable cytochrome P450 (CYP175A1) cloned from Thermus thermophilus shows mid-point unfolding temperature (Tm) of 88 °C (361 K) along with high thermodynamic stability making it a potential industrially viable biocatalyst. Molecular docking analyses, and structural superposition with steroidogenic and fatty acid metabolizing cytochrome P450 s suggested that the tyrosine 68 may have important role in binding as well as metabolism of substrates by the enzyme. Site-saturation mutation of the tyrosine 68 residue was carried out and several unique mutations were obtained that were properly folded and showed high thermostability. We investigated the effects of variation of the single residue, Tyr68 at the substrate binding pocket of the enzyme on the substrate specificity of CYP175A1. Screening of the mutant colonies of CYP175A1 obtained after saturation mutagenesis of Tyr68 using saturated fatty acid, myristic acid and poly unsaturated fatty acids showed that the Y68K had notable binding and catalytic activity for mono-oxygenation of the saturated fatty acid (myristic acid), which had no major detectable binding affinity towards the WT enzyme. The Y68R mutant of CYP175A1, on the other hand was found to selectively bind and catalyse reaction of cholesterol. The wild type as well as both the mutants of the enzyme however bind poly unsaturated fatty acids. The results thus show that saturation mutation of a single amino acid at the substrate binding pocket of the thermostable cytochrome P450 could induce sufficient changes in the substrate binding pocket of the enzyme that can efficiently change substrate specificity of the enzyme.

The effect of the mutation at the core of the ferritin nanocage (apo‐rHLFr) on the uptake of IrCp* has been investigated by structural and spectroscopic methods. Site‐specific mutations of two polar residues viz., Asp38 and Arg52 were investigated. The uptake of IrCp* was increased by about 1.5‐fold on mutation of Arg52 by His compared to the wild‐type variant, while mutation of Asp38 by His had no effect on the uptake. All the variants of the Ir‐embedded ferritin cages remained as stable as the wild‐type analogue. These hybrid bio‐nanocages of ferritin were found to efficiently catalyze transfer hydrogenation of various substituted acetophenones forming the corresponding chiral alcohols with up to 88 % conversion and 70 % enantioselectivity. An electron‐withdrawing substituent on the reactant enhanced the Turnover frequency of the reaction. Molecular docking analyses suggested that the substrate binds in different orientations at the active site in different mutants of the nanocage.