Alterations In Secondary Structure Research Articles

Comparative Study of Time-Dependent Aluminum Exposure and Post-Exposure Recovery Shows Better Improvement in Synaptic Changes andNeuronal Pathology in Rat Brain After Short-Term Exposure.

Aluminum is a ubiquitous metal that causes multiple brain pathologies such as, cognitive dysfunction and Alzheimer's disease like symptoms. Exposure to aluminum through drinking water is responsible for hampering learning and memory. This study aimed to compare (1) the time-dependent effect of aluminum exposure (keeping total exposure of 5850mg/kg same) in two durations, 30 and 45days, and (2) to compare post-exposure self-recovery effect after 20days in both (30 and 45days exposure) groups. Rats were given 130 and 195mg/kg of AlCl3·6H2O for 45 and 30days respectively, to see the time-dependent exposure effect. At the end of exposure, rats were given distilled water and allowed to self-recover for 20days to study the recovery. Expressionlevelsof synaptic genes (Syp, SNAP25, Nrxn1/2, PSD95, Shank1/2, Homer1, CamkIV, Nrg1/2 and Kalrn) were measured using qPCR and compared in the exposure and recovery groups. Cellular morphology of the rat brain cortex and hippocampus was also investigated. Damage in lipid and protein profile was measured by employing FTIR. Results showed downregulation of mRNA expression of synaptic genes, plaques deposition, disorganization in lipid and protein profile by increasing membrane fluidity, and disorder and alteration of protein secondary structure after both exposure periods. However, better improvement/recovery in these parameters were observed in recovery group of 30days aluminum exposure compared to 45days aluminum exposure group. Taken together, these results suggested that short-term exposure resulted in better restoration of lipid and protein profile after time-dependent exposure of aluminum than prolonged exposure.

Evaluation of copper-induced biomolecular changes in different porin mutants of Escherichia coli W3110 by infrared spectroscopy.

Copper (Cu), one of the heavy metals, plays a vital role in many complex biochemical reactions as a trace element. However, it often becomes toxic when its concentration in the cell exceeds a certain level. Homeostasis of metals in the cell is primarily related to regulating metal transport into and out of the cell. Therefore, it is thought that porin proteins, which have a role in membrane permeability, may also play a role in developing Cu resistance. This study identified the differences between the molecular profiles of wild-type Escherichia coli W3110 and its seven different porin mutants exposed to Cu ions using attenuated total reflectance (ATR)-Fourier transform infrared (FTIR) spectroscopy. The results showed that the absence of porin genes elicits global changes in the structure and composition of membrane lipids and proteins, in both the absence and presence of Cu. The lack of porin genes significantly elevated the amounts of fatty acids and phospholipids. When the alterations in protein secondary structures were compared, the quantity of amide I proteins was diminished by the presence of Cu. However, the amount of amide II proteins increased in porin mutant groups independent of Cu presence or absence. The DNAs are transformed from B- and Z-form to A-form due to porin mutations and the presence of Cu ions. The lack of porin genes increased polysaccharide content independent of Cu presence. This study can help characterize Cu detoxification efficiency and guide for obtaining active living cells to be used in bioremediation.

Improving the Functionality of Lentil–Casein Protein Complexes through Structural Interactions and Water Kefir-Assisted Fermentation

Highly nutritious lentil proteins (LP) have recently attracted interest in the food industry. However, due to their low solubility, extensive application of LP is severely limited. This study describes a new and successful method for overcoming this challenge by improving the nutritional–functional properties of LP, particularly their solubility and protein quality. By combining protein complexation with water kefir-assisted fermentation, the water solubility of native LP (~58%) increases to over 86% upon the formation of lentil–casein protein complexes (LCPC). Meanwhile, the surface charge increases to over −40 mV, accompanied by alterations in secondary and tertiary structures, as shown by Fourier-transform infrared and UV-vis spectra, respectively. In addition, subjecting the novel LCPC to fermentation increases the protein digestibility from 76% to over 86%, due to the reduction in micronutrients that have some degree of restriction with respect to protein digestibility. This approach could be an effective and practical way of altering plant-based proteins.

Resilience study of wheat protein networks with large amplitude oscillatory shear rheology

This study investigates the effect of standard laboratory production steps on the network properties of vital gluten. Gluten experiences high mechanical input during production from wheat dough and subsequent processing, which influences the network characteristics. The production of vital gluten in laboratory scale ensures full control on all processing parameters and the possibility to obtain the protein using the mildest washing, drying and milling techniques. Large Amplitude Oscillatory Shear (LAOS) tests were performed on wheat dough, intermediate materials during gluten production and the final product vital gluten at frequencies of 1–15 rad/s and strains from 0.01 to 100%. The elastic and viscous behavior of the material are described by analyzing the fundamental response and the higher harmonics, based on Lissajous-Bowditch curves. The intra-cycle behavior revealed that vital gluten only exhibits network stiffening, whereas dough shows both strain stiffening and shear thinning effects. This suggests that starch induces the shear thinning effects in dough.Comparison of rehydrated and fresh gluten obtained from the same starting material reveals minor alteration of the protein secondary structures during the production process by means of Fourier-transform infrared spectroscopy (FTIR). A pronounced difference in water uptake accounts for modified rheological network properties.

Histopathology of Corneal Lenticules Obtained from Small Incision Lenticule Extraction (SMILE) versus Microkeratome Excision.

To study the alterations on the lenticules extracted after femtosecond (Femto) small incision lenticule extraction (SMILE) versus the corneal free cap removed using a microkeratome. The visuMax (500 kHz; laser energy: 180 nJ) was used for small-incision lenticule extraction. Free caps from human cadaveric corneas were excised by microkeratome. The collected lenticules were examined with the light and transmission electron microscope (TEM) for histological analysis, DNA fragmentation was assessed by agarose gel electrophoresis, DNA damage was evaluated using comet assay, and corneal proteins secondary structure was assessed by Fourier transform infrared spectroscopy (FTIR). Light microscopic examination showed the presence of more edematous stroma under Femto SMILE than under free cap with a percentage change of 101.6%. In the Femto SMILE group, TEM examination showed pyknotic keratocytes, disruption, and cavitation of the collagen arrays stromal area under Femto SMILE. The DNA fragmentation for the Femto SMILE group revealed one undefined band with a size of 1.1 Kbp. The comet assay analysis indicated the presence of 3% and 8.0% tailed cells for the free cap and Femto SMILE groups, respectively. The tail lengths were 1.33 0.16 and 1.67 0.13 µm (P 0.01), the percentage of tail DNA was 1.41 0.18% (P 0.01) and 1.72 0.15%, and the tail moments were 1.88 0.12 AU and 2.87 0.14 AU (P 0.001) for the free cap and Femto SMILE groups, respectively. FTIR spectroscopy of the Femto smile group revealed disorders in the secondary and tertiary structure of the proteins. Femto SMILE technique induced more structural changes, DNA fragmentation, DNA damage, and corneal proteins secondary structure alteration than those induced by a microkeratome cutting. These changes may be attributed to the deep penetration of high energy levels to the corneal layer. These findings may highlight the potential impact of the Femto SMILE on the cornea and the necessity for managing the laser parameters used.

Effect of starch on the gel properties and protein conformation of egg white subjected to alkali‐heat treatment

SummaryThe colour, texture, water‐holding capacity (WHC) and particle properties of gels were examined to assess the impact of starch (cassava, corn and sweet potato starch) on the attributes of alkali‐heat‐induced gels. The results indicated that the whiteness, hardness and WHC of egg white gels were positively affected by starches. The microstructure of the mixed gel showed increasing reticulation and holes as the starch content increased, which contributed to the stiff structure of the gel. Although the gel made with corn starch had the lowest WHC, the gel made with sweet potato starch had the highest WHC. The average size of the gel particles decreased as a result of the inhibition of starch gelatinisation. Starch causes alterations in the protein secondary structure of egg white gels, primarily causing α‐helix and β‐sheet interconversion. The stiffness of the gels was improved by increasing the quantity of β‐sheet structures. According to the electrophoresis findings, a small amount of sweet potato starch can help prevent ovalbumin from degrading, while having no impact on the ability of the protein to cross‐link.

Structural effects of ALS-associated mutations on profilin.

Amyotrophic Lateral Sclerosis (ALS), known as Lou Gehrig's disease, which has been on the rise (approximately 31000 patients in the USA, according to CDC), has been associated with specific mutations on Profilin, a protein with distinct binding sites for both poly-L-proline (PLP) and Actin. Here in this computational study, we have investigated the effect of a few of these mutations inducing conformational differences and alteration of secondary structures that could be the molecular reason behind the ALS. Among a few potential mutations associated with the disease, we have investigated the effect of G118V and T109M mutations and their combination in the conformational changes of Profilin and Actin in the presence of PLP. Using close to 30 microseconds of all-atom molecular dynamics simulations across 16 systems of study composed of Profilin, Actin, and PLP, we have identified intermolecular interactions that could be among the main reasons for conformational differences between wild-type Profilin and mutants and potentially the reasons for the ALS in the molecular level due to the mutations. Our findings help to gain a better molecular-level understanding of ALS. We can use the plethora of structural and conformational information and intermolecular interactions to design novel therapeutics for the disease.

Surface Ligand Influences the Cu Nanoclusters as a Dual Sensing Optical Probe for Localized pH Environment and Fluoride Ion.

Functional metal nanomaterials, especially in the nanocluster (NC) size regime, with strong fluorescence, aqueous colloidal stability, and low toxicity, necessitate their application potential in biology and environmental science. Here, we successfully report a simple cost-effective method for red-/green-color-emitting protein/amino-acid-mediated Cu NCs in an aqueous medium. As-synthesized Cu NCs were characterized through UV-Vis absorption spectroscopy, fluorescence spectroscopy, time-resolved photoluminescence, dynamic light scattering, zeta potential, transmission electron microscopy and X-ray photoelectron spectroscopy. The optical properties of both Cu NCs responded linearly to the variation in pH in the neutral and alkaline ranges, and a robust pH reversible nature (between pH 7 and 11) was observed that could be extended to rapid, localized pH sensor development. However, a contrasting pH response nature between protein-Cu NCs and amino acid-Cu NCs was recorded. The alteration in protein secondary structure and strong binding nature of the surfactants were suggested to explain this behavior. Furthermore, we investigated their use as an efficient optical probe for fluoride ion detection. The limit of detection for protein-Cu NCs is 6.74 µM, whereas the limit of detection for amino acid-Cu NCs is 4.67 µM. Thus, it is anticipated that ultrasmall Cu NCs will exhibit promise in biological and environmental sensing applications.

RNA Secondary Structure Alteration Caused by Single Nucleotide Variants.

A point mutation in coding RNA can cause not only an amino acid substitution but also a dynamic change of RNA secondary structure, leading to a dysfunctional RNA. Although in silico structure prediction has been used to detect structure-disrupting point mutations known as riboSNitches, exhaustive simulation of long RNAs is needed to detect a significant enrichment or depletion of riboSNitches in functional RNAs. Here, we have developed a novel algorithm Radiam (RNA secondary structure Analysis with Deletion, Insertion, And substitution Mutations) for a comprehensive riboSNitch analysis of long RNAs. Radiam is based on the ParasoR framework, which efficiently computes local RNA secondary structures for long RNAs. ParasoR can compute a variety of structure scores over globally consistent structures with maximal span constraints for the base pair distance. Using the reusable structure database made by ParasoR, Radiam performs an efficient recomputation of the secondary structures for mutated sequences. An exhaustive simulation of Radiam is expected to find reliable riboSNitch candidates on long RNAs by evaluating their statistical significance in terms of the change of local structure stability.

Effects of alternating electric field assisted freezing-thawing-aging sequence on longissimus dorsi muscle microstructure and protein characteristics

The current study investigates the influence of alternating electric field (AEF)-assisted freezing-thawing-aging sequence on the muscle microstructure and myofibrillar protein characteristics. Three treatments were used for longissimus dorsi (LD) muscle: only aging (OA), freezing-thawing-aging sequence (FA) and AEF-assisted freezing-thawing-aging sequence (EA). Compared with the FA and EA groups, the OA group showed considerably fewer cracks between muscle fibers and maintained the integrity of the Z-line as observed using scanning and transmission electron microscopy, respectively. Furthermore, the EA treatment effectively decreased myofibrillar fragmentation, myofibrillar protein aggregation, and protein oxidation, as shown by the myofibrillar fragmentation index, turbidity, and total sulfhydryl concentration. Analysis of surface hydrophobicity and the Fourier transform infrared, UV absorption, and fluorescence spectrums indicated that AEF minimized the alterations of protein secondary and tertiary structure alterations during aging after freezing.

Glycation restrains open-closed conformation of Insulin

In hyperglycemic conditions, the level of reactive dicarbonyl metabolites concentration is found to be high, which plays a significant role in protein glycation. Despite decades of research, the effect of methylglyoxal on the structure and function of insulin is still unknown. Through a shift in conformation at the B-chain C-terminal (BT-CT) hinge from an "open" to a "wide-open" conformation, insulin binds to the receptor and activates the signal cascade. Insulin resistance, which is the main sign of Type 2 Diabetes, can be caused by a lack of insulin signaling. Methylglyoxal site-specific glycation in residue R22 at B chain forms AGE product Methylglyoxal-hydroimidazolone (MGH1) in insulin. In this work, we present molecular dynamics study of this glycated insulin R22MGH1, which revealed new insights into the conformational and structural changes. We find the following key results: 1) B-chain in insulin undergoes a closed conformational change upon glycation. 2) Glycated insulin shows secondary structure alteration. 3) Glycated insulin retains its closed shape due to an unusually strong hydrophobic contact between B-chain residues. 4) Wide open native conformation of insulin allows the B chain helix to be surrounded by more water molecules compared to the closed conformation of glycated insulin. The closed conformation of glycated insulin impairs its binding to insulin receptor (IR).

In vitro and in vivo immunogenicity assessment of protein aggregate characteristics

The immunogenicity risk of therapeutic protein aggregates has been extensively investigated over the past decades. While it is established that not all aggregates are equally immunogenic, the specific aggregate characteristics, which are most likely to induce an immune response, remain ambiguous. The aim of this study was to perform comprehensive in vitro and in vivo immunogenicity assessment of human insulin aggregates varying in size, structure and chemical modifications, while keeping other morphological characteristics constant. We found that flexible aggregates with highly altered secondary structure were most immunogenic in all setups, while compact aggregates with native-like structure were found to be immunogenic primarily in vivo. Moreover, sub-visible (1–100 µm) aggregates were found to be more immunogenic than sub-micron (0.1–1 µm) aggregates, while chemical modifications (deamidation, ethylation and covalent dimers) were not found to have any measurable impact on immunogenicity. The findings highlight the importance of utilizing aggregates varying in few characteristics for assessment of immunogenicity risk of specific morphological features and may provide a workflow for reliable particle analysis in biotherapeutics.

Pb(II) coordination to the nonclassical zinc finger tristetraprolin: retained function with an altered fold.

Tristetraprolin (TTP) is a nonclassical CCCH zinc finger (ZF) that plays a crucial role in regulating inflammation. TTP regulates cytokine mRNAs by specific binding of its two conserved ZF domains (CysX8CysX5CysX3His) to adenylate-uridylate-rich sequences (AREs) at the 3'-untranslated region, leading to degradation of the RNA. Dysregulation of TTP in animal models has demonstrated several cytokine-related syndromes, including chronic inflammation and autoimmune disorders. Exposure to Pb(II), a prevalent environmental toxin, is known to contribute to similar pathologies, in part by disruption of and/or competition with cysteine-rich metalloproteins. TTP's role during stress as a ubiquitous translational regulator of cell signaling (and dysfunction), which may underpin various phenotypes of Pb(II) toxicity, highlights the importance of understanding the interaction between TTP and Pb(II). The impact of Pb(II) binding on TTP's fold and RNA-binding function was analyzed via UV-Vis spectroscopy, circular dichroism, X-ray absorption spectroscopy, nuclear magnetic resonance spectroscopy, and fluorescence anisotropy. A construct containing the two ZF domains of TTP (TTP-2D) bound to Pb(II) with nanomolar affinity and exhibited a different geometry and fold in comparison to Zn2-TTP-2D. Despite the altered secondary structure, Pb(II)-substituted TTP-2D bound a canonical ARE sequence more selectively than Zn2-TTP-2D. Taken together, these data suggest that Pb(II) may interfere with proper TTP regulation and hinder the cell's ability to respond to inflammation.

Dydrogesterone disrupts lipid metabolism in zebrafish brain: A study based on metabolomics and Fourier transform infrared spectroscopy

Brain is a potential target for neuroprogestogens and/or peripheral progestogens. Previous studies reported that expression of genes about steroidogenesis, reproduction, cell cycle, and circadian rhythm in zebrafish brain could be affected by progestogens. However, there are limited information from metabolites or biomacromolecules aspects, leaving an enormous gap in understanding toxic effects of progestogens on fish brain. In this study, we exposed zebrafish embryos to 2.8, 27.6, and 289.8 ng/L dydrogesterone (DDG, a synthetic progestogen) until sexual maturity (140 days). LC-MS and GC-MS based untargeted metabolomics and Fourier-transform infrared (FTIR) spectroscopy were then performed to investigate the metabolic profiles and macromolecular changes of brain of these zebrafish. The results from multivariate statistical analysis of metabolite features showed a clear separation between different treatment groups of both female and male zebrafish brains. DDG exposure increased the levels of cholesterol, saturated fatty acids, and nucleoside monophosphates, but decreased the contents of polyunsaturated fatty acids (PUFAs), lysophosphatides, and nucleosides in dose-dependent manner. FTIR results indicated that DDG exposure led to accumulation of saturated lipids, reduction of nucleic acids and carbohydrates, and alteration of protein secondary structures. The findings from this study demonstrated that DDG could affect contents of metabolites and biomacromolecules of zebrafish brain, which may finally lead to brain dysfunctions.

Exploring the mechanism of compromised thermostability of aromatic l-amino acid decarboxylase from Bacillus atrophaeus through comparative molecular dynamics simulations

As an excellent biocatalyst, aromatic l-amino acid decarboxylase from Bacillus atrophaeus (BaAADC) catalyzes the production of various vital aromatic biogenic amines. However, its weak heat resistance makes BaAADC unsuitable for industrial production. Here, we generated the three-dimensional structure of BaAADC and used molecular dynamics simulations to investigate the dynamic behavior of BaAADC at different temperatures (303–343 K) to investigate the association between its thermal stability and internal residues. We have shown that the natural structure of BaAADC is altered at high temperatures, with more striking alterations in secondary structure at the active site, fewer internal hydrogen bonding, and greater amino acid flexibility in the active pocket. Based on the root mean square fluctuations, the residues ASN46, GLU54, ALA113, ASP324, VAL329, and GLU332 were confirmed to be the most flexible. Salt bridge analysis revealed ion pairs ASP182-ARG197, ASP299-ARG347, GLU121-ARG344, GLU326-ARG316, GLU337-ARG316, GLU341-ARG347, and GLU470-ARG427 were partially responsible for the inactivation of BaAADC at high temperatures. In addition, strategies to improve the thermostability of BaAADC were discussed. This research might provide light on the development and modification of thermostable BaAADC.

Characterization of oyster water-soluble protein-EGCG conjugate and its antioxidant effects on linolic acid in emulsion system

(−)-Epigallocatechin-3-gallate (EGCG) is frequently used as a good antioxidant in food systems. In contrast, its antioxidant capacity was limited in the oil-in-water emulsion system due to its high hydrophilicity. Therefore, food-derived proteins with higher hydrophobicity were utilized to bind with EGCG and form new molecules that could stay in the oil-water interface to improve the stability of such emulsions. This study combined water-soluble protein (OWPs) extracted from oysters with EGCG to form the OWPs-EGCG complex. When combined with EGCG, the molecular weight and particle diameter (nearly 180–240 nm) of OWPs slightly increased in a dose-dependent manner by increasing the proportion of EGCG. Moreover, several conformational changes to OWPs by EGCG addition were found, such as the alteration of secondary structures, the fluorescence quenching, and the involvement of binding groups. The OWPs-EGCG complex (1:0.025) exhibited the highest emulsifying activities, while the ratio of 1:0.2 had the highest foaming stability. Furthermore, the OWPs-EGCG complex showed nearly 90% free radical scavenging activities with an increasing ratio of EGCG. In the established temporary storage model of OWPs-EGCG conjugate/linolic acid emulsion system, the OWPs-EGCG complex exhibited enhanced antioxidant effects against the oxidation of OWPs and linolic acid. This study provided a theoretical basis for the potential applications of food-derived proteins/EGCG conjugates on antioxidant effects against lipid oxidation in the emulsifying system.

The Hidden Role of Non-Canonical Amyloid β Isoforms in Alzheimer's Disease.

Recent advances have placed the pro-inflammatory activity of amyloid β (Aβ) on microglia cells as the focus of research on Alzheimer's Disease (AD). Researchers are confronted with an astonishing spectrum of over 100 different Aβ variants with variable length and chemical modifications. With the exception of Aβ1-42 and Aβ1-40, the biological significance of most peptides for AD is as yet insufficiently understood. We therefore aim to provide a comprehensive overview of the contributions of these neglected Aβ variants to microglia activation. First, the impact of Aβ receptors, signaling cascades, scavenger mechanisms, and genetic variations on the physiological responses towards various Aβ species is described. Furthermore, we discuss the importance of different types of amyloid precursor protein processing for the generation of these Aβ variants in microglia, astrocytes, oligodendrocytes, and neurons, and highlight how alterations in secondary structures and oligomerization affect Aβ neurotoxicity. In sum, the data indicate that gene polymorphisms in Aβ-driven signaling pathways in combination with the production and activity of different Aβ variants might be crucial factors for the initiation and progression of different forms of AD. A deeper assessment of their interplay with glial cells may pave the way towards novel therapeutic strategies for individualized medicine.

Secondary structure alterations of RBC assessed by FTIR-ATR in correlation to 2,3-DPG levels in ApoE/LDLR–/– Mice

In the present study, we characterized the secondary structure alterations of intact red blood cells (RBCs) cytosol with special attention to the sex-related alterations in 8- and 24-week-old female and male ApoE/LDLR-/- mice, compared to age-matched female and male C57BL/6J control animals. Results were obtained with previously established methodology based on Fourier transform infrared spectroscopy-attenuated total reflectance (FTIR-ATR). Additionally, we evaluated 2,3-DPG levels in the RBCs and showed its potential link to the hemoglobin (Hb) secondary structure alterations. Considering Hb structure alterations probed by FTIR-ATR, the ratio of turns to α–helices in 8-week-old ApoE/LDLR-/- mice suggested more pronounced secondary structure alterations within the RBCs than in the age-matched control. Sex-related differences were observed solely in 24-week-old male ApoE/LDLR-/- mice, which showed statistically significant increase in the secondary structure alterations compared to 24-week-old female ApoE/LDLR-/- mice. Similar to the secondary structure alterations, no sex-related differences were observed in the levels of 2,3-DPG in RBCs, except for 24-week-old male ApoE/LDLR-/- mice, which showed significantly higher levels compared to the age-matched female ApoE/LDLR-/- mice. Considering the age-related alterations, we observed significant increases in the intracellular 2,3-DPG of RBCs with animals’ age in all studied groups, except for female ApoE/LDLR-/- mice, where a significant difference was not reported. This suggests the clear correlation between secondary structure of Hb alterations and 2,3-DPG levels for male and female murine RBC and proves a higher resistance of older female RBCs to the secondary structure changes with progression of atherosclerosis. Moreover, it may be concluded that higher 2,3-DPG levels in RBCs occurred in response to the secondary structure alterations of Hb in ApoE/LDLR-/- mice.

Application of ultrasound treatment to improve the technofunctional properties of hemp protein isolate

The hemp seed protein isolate (HPI) was extracted from defatted hemp seed meal using alkaline extraction (pH = 10.0) acid precipitation (pH = 4.3) method. The effects of ultrasound treatment on its solubility, emulsifying properties, particle size, microstructure, molecular subunits, secondary and tertiary structures, and surface hydrophobicity were determined. Solubility and emulsifying properties of HPI were significantly improved after ultrasound treatment (20 kHz) at 400 W for 12 min. The solubility, emulsion activity, and emulsion stability indices increased by 3.7- 1.7- and 1.5-fold, respectively. The improvement in functional properties correlated with reduced particle size, increased surface hydrophobicity and the alteration in secondary and tertiary structure of HPI. The solubility and emulsifying properties of the ultrasound treated HPI were better than native pea protein isolate but still not as good as those of native soy protein isolate for which further research is needed.

Mechanistic study and synergistic effect on inhibition of α-amylase by structurally similar flavonoids

Natural plant-based flavonoids have the potential effect of reducing postprandial hyperglycaemia, thus they can be used to prevent and control type 2 diabetes mellitus. Here, to explore the antihyperglycemic mechanisms of flavonoids (apigenin, scutellarein, hispidulin and nepetin), their inhibition mechanisms against α-amylase were evaluated using multispectral methods and molecular docking in this study. As a result, nepetin, a competitive inhibitor, exhibited the best inhibitory effect than other three flavonoids, suggesting that adjacent dihydroxyl group on the B-ring played an important role in inhibiting the activity of α-amylase. At the same time, the combination of apigenin and scutellarein/hispidulin/nepetin synergistically improved their inhibitory effects on α-amylase. In addition, fluorescence quenching analysis indicated that the addition of flavonoids could result in the static quenching of α-amylase and the formation of non-covalent complexes. The interaction further led to the alteration of microenvironment and secondary structure of α-amylase, thus reducing the catalytic activity of enzyme. In conclusion, this finding can provide vital insights into the use of flavonoids, especially nepetin, as the potential drug candidates for the treatment of diabetes.