Characterization of physicochemical and immunogenic properties of allergenic proteins altered by food processing:a review

Food allergens are mainly naturally-occurring proteins with immunoglobulin E(IgE)-binding epitopes.Understanding the structural and immunogenic characteristics of allergenic proteins is essential in assessing whether and how food processing techniques reduce allergenicity.We here discuss the impacts of food processing technologies on the modification of physicochemical,structural,and immunogenic properties of allergenic proteins.Detection techniques for characterizing changes in these properties of food allergens are summarized.Food processing helps to reduce allergenicity by aggregating or denaturing proteins,which masks,modifies,or destroys antigenic epitopes,whereas,it cannot eliminate allergenicity completely,and sometimes even improves allergenicity by exposing new epitopes.Moreover,most food processing techniques have been tested on purified food allergens rather than food products due to potential interference of other food components.We provide guidance for further development of processing operations that can decrease the allergenicity of allergenic food proteins without negatively impacting the nutritional profile.

Grouping of amino acids and recognition of protein structurally conserved regions by reduced alphabets of amino acids

Sequence alignment is a common method for finding protein structurally conserved/similar regions. However, sequence alignment is often not accurate if sequence identities between to-be-aligned se- quences are less than 30%. This is because that for these sequences, different residues may play similar structural roles and they are incorrectly aligned during the sequence alignment using substitu- tion matrix consisting of 20 types of residues. Based on the similarity of physicochemical features, residues can be clustered into a few groups. Using such simplified alphabets, the complexity of protein sequences is reduced and at the same time the key information encoded in the sequences remains. As a result, the accuracy of sequence alignment might be improved if the residues are properly clustered. Here, by using a database of aligned protein structures (DAPS), a new clustering method based on the substitution scores is proposed for the grouping of residues, and substitution matrices of residues at different levels of simplification are constructed. The validity of the reduced alphabets is confirmed by relative entropy analysis. The reduced alphabets are applied to recognition of protein structurally conserved/similar regions by sequence alignment. The results indicate that the accuracy or efficiency of sequence alignment can be improved with the optimal reduced alphabet with N around 9.

Structural changes of proteins in liver cirrhosis and consequential changes in their function

The liver is the site of synthesis of the majority of circulating proteins.Besides initial polypeptide synthesis,sophisticated machinery is involved in the further processing of proteins by removing parts of them and/or adding functional groups and small molecules tailoring the final molecule to suit its physiological purpose.Posttranslational modifications(PTMs)design a network of molecules with the common protein ancestor but with slightly or considerably varying activity/localization/purpose.PTMs can change under pathological conditions,giving rise to aberrant or overmodified proteins.Undesired changes in the structure of proteins most often accompany undesired changes in their function,such as reduced activity or the appearance of new effects.Proper protein processing is essential for the reactions in living beings and crucial for the overall quality control.Modifications that occur on proteins synthesized in the liver whose PTMs are cirrhosis-related are oxidation,nitration,glycosylation,acetylation,and ubiquitination.Some of them predominantly affect proteins that remain in liver cells,whereas others predominantly occur on proteins that leave the liver or originate from other tissues and perform their function in the circulation.Altered PTMs of certain proteins are potential candidates as biomarkers of liver-related diseases,including cirrhosis.This review will focus on PTMs on proteins whose structural changes in cirrhosis exert or are suspected to exert the most serious functional consequences.

Prediction of Protein Structural Classes Using the Theory of Increment of Diversity and Support Vector Machine

Based on the concept of the pseudo amino acid composition （PseAAC）, protein structural classes are predicted by using an approach of increment of diversity combined with support vector machine （ID-SVM）, in which the dipeptide amino acid composition of proteins is used as the source of diversity. Jackknife test shows that total prediction accuracy is 96.6% and higher than that given by other approaches. Besides, the specificity （Sp） and the Matthew＇s correlation coefficient （MCC） are also calculated for each protein structural class, the Sp is more than 88%, the MCC is higher than 92%, and the higher MCC and Sp imply that it is credible to use ID-SVM model predicting protein structural class. The results indicate that： 1 the choice of the source of diversity is reasonable, 2 the predictive performance of IDSVM is excellent, and3 the amino acid sequences of proteins contain information of protein structural classes.

Changes of structure and functional properties of rice protein in the fresh edible rice during the seed development

It has been reported that fresh edible rice has more bioactive compounds and its protein is easier to digest and has lower hypoallergenic than mature rice. In this paper, the changes in structure and functional properties of proteins at five different stages, including early milky stage(EMS), middle milky stage(MMS), late milky stage(LMS), waxy ripe stage(WS)and ripening stage(RS), during the seed development were investigated. It was found that with the seed developing, the molecular weight of fresh rice protein gradually become larger while the secondary structure changed from the highest content of disordered structure at MMS to the highest content of ordered structure at RS, which affect the surface hydrophobicity and then the functional properties of proteins, including foaming properties, emulsifying properties and oil holding capacity. Fresh rice protein at MMS has the strongest surface hydrophobicity while fresh edible rice protein at RS has the strongest oil holding capability. The results of our study can provide a theoretical basis for the application of fresh rice protein in the food industry and help to develop new fresh edible rice food.

Structural modeling of proteins by integrating small-angle x-ray scattering data

Elucidating the structure of large biomolecules such as multi-domain proteins or protein complexes is challenging due to their high flexibility in solution. Recently, an ＂integrative structural biology＂ approach has been proposed, which aims to determine the protein structure and characterize protein flexibility by combining complementary high- and lowresolution experimental data using computer simulations. Small-angle x-ray scattering（SAXS） is an efficient technique that can yield low-resolution structural information, including protein size and shape. Here, we review computational methods that integrate SAXS with other experimental datasets for structural modeling. Finally, we provide a case study of determination of the structure of a protein complex formed between the tandem SH3 domains in c-Cb1-associated protein and the proline-rich loop in human vinculin.

Bioinformatic Analysis of Structural Proteins of Paramyxovirus Tianjin Strain

The amino acid sequences of the NP, P, M, F, HN and L proteins of the paramyxovirus Tianjin strain were analyzed by using the bioinformatics methods. Phylogenetic analysis based on 6 structural proteins among the Tianjin strain and 25 paramyxoviruses showed that the Tianjin strain belonged to the genus Respirovirus, in the subfamily Paramyxovirinae, and was most closely related to Sendai virus (SeV). Phylogenetic analysis with 14 known SeVs showed that Tianjin strain represented a new evolutionary lineage. Similarities comparisons indicated that Tianjin strain P protein was poorly conserved, sharing only 78.7% - 91.9% amino acid identity with the known SeVs, while the L protein was the most conserved, having 96.0% - 98.0% amino acid identity with the known SeVs. Alignments of amino acid sequences of 6 structural proteins clearly showed that Tianjin strain possessed many unique amino acid substitutions in their protein sequences, 15 in NP, 29 in P, 6 in M, 13 in F, 18 in HN, and 29 in L. These results revealed that Tianjin strain was most likely a new genotype of SeV. The presence of unique amino acid substitutions suggests that Tianjin strain maybe has a significant difference in biological, pathological, immunological, or epidemiological characteristics from the known SeVs.

The Structural Proteins of Bdellovibrio bacteriovorus Bacteriophage MAC-1

In the present investigation the structural proteins associated with MAC-1 bacteriophage have been characterized using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE);tandem mass spectrometry of protein bands from SDS-PAGE gel;from the open reading frames (ORFs) deduced from MAC-1 genome sequence and amino acid sequence homology searches from the Uniprot database (up000002418). Results have led to the identification of at least three structural proteins associated with MAC-1 phage genome. They are: capsid protein (~55,000-daltons);spike protein (~22,000-daltons) and a low molecular weight DNA binding protein (~4000-dal- tons). In addition, two other minor proteins were tentatively identified as replicative and scaffold proteins based on two to three unique peptides from mass spectrometry data. However, other proteins coded (ORFs) by phage genome remain to be identified.

Prokaryotic Expression and Immunogenicity of Dominant Epitope Region of Goose Parvovirus Structural Protein VP3

[Objective] The paper was to develop a subunit vaccine candidate for prevention and control of goose parvovirus infection. [Method]Based on the prokaryotic expression system, the antigenic epitopes and locations of the structural protein VP3 were predicted by software analysis,and the region displaying a large portion of antigenic epitopes was amplified by PCR. The target VP3 DNA fragment was inserted into pET-30 a-VP3 vector, was transformed into Escherichia coli BL21 competent cells for protein expression and animal test. The SPF chickens were immunized with the recombinant protein and the antisera were collected for neutralization test by using a goose embryo fibroblast. [Result] The recombinant plasmid was constructed, and the target region of VP3 protein was expressed efficiently in a soluble form. The neutralizing titers of antisera could reach up to-2.608. [Conclusion] The target region displaying a large portion of antigenic epitopes of the structural protein VP3 could be expressed efficiently in soluble form, and the expressed protein could induce neutralizing antibodies in SFP chicken.

A Novel Design Method for Protein-Like Molecules from the Perspective of Sheaf Theory

Proteins perform a variety of functions in living organisms and their functions are largely determined by their shape. In this paper, we propose a novel mathematical method for designing protein-like molecules of a given shape. In the mathematical model, molecules are represented as loops of n-simplices (2-simplices are triangles and 3-simplices are tetrahedra). We design a new molecule of a given shape by patching together a set of smaller molecules that cover the shape. The covering set of small molecules is defined using a binary relation between sets of molecules. A new molecule is then obtained as a sum of the smaller molecules, where addition of molecules is defined using transformations acting on a set of (n + 1)-dimensional cones. Due to page limitations, only the two-dimensional case (i.e., loops of triangles) is considered. No prior knowledge of Sheaf Theory, Category Theory, or Protein Science is required. The author hopes that this paper will encourage further collaboration between Mathematics and Protein Science.

Quantum Chemical Studies on Proteins in the Reaction Center of Rhodobacter sphaeroides

The electronic structure of protein chains L and M in photosynthetic reaction center (PRC) of Rhodobacter sphaeroides (Van Niel) Imhoff, Truper et Pfennig) was studied by using the Overlapping Dimer Approximation method and the Extended Negative Factor Counter method at ab initio level. The result indicated that: (1) Amino acid residues, the molecular orbitals of which composed the main components of frontier orbitals of protein chain L (M), are located at the random coil areas of chain L (alpha helix areas of chain M). Since the random coil is flexible and more easy to change its conformation in the electron transfer process and to reduce the energy of the system, and the structure of the alpha helix is reletively stable, this difference might be one of the causes for the electron transfer in photosynthetic reaction center (PRC) only takes place along the L branch. (2) The His residues which axially coordinated to the 'special pair' P and accessory chlorophyll molecules (ABChls) are essentially important for the E-LUMO levels of P and ABChl. But, the corresponding molecular orbitals of these His residues do not appear in the composition of frontier orbitals of protein chains. It means that the interaction between pigment molecules and protein chains do not influence the contribution to the frontier orbitals of protein chains explicitly, but influences the corresponding E-LUMO levels significantly.

Changes in physicochemical characteristics of wheat flour and quality of fresh wet noodles induced by microwave treatment

Fresh wet noodles(FWN) are popular staple foods due to its unique chewy texture and favorable taste. However,the development of FWN is limited by its short shelf life and high browning rate. It has been found that the quantity of original microorganisms in wheat flour produced by traditional method is relatively high, which is detrimental to the processing quality and storage stability of FWN. Consequently, it becomes imperative to decrease microorganisms in wheat flour. Microwave treatment has been regarded as a promising method in the food industry due to its potential in inhibiting microbial growth and inactivating enzymes without causing adverse effect on the food quality. This study aims to investigate the effects of microwave treatment of wheat kernels under different powers(1, 2, 3, 4, 5 kW) on the physicochemical properties of wheat flour and the quality of FWN. The results revealed that microwave treatment had a significant effect on microbial inhibition and enzyme inactivation, wherein the total plate count(TPC) and yeast and mold counts(YMC) decreased by 0.87 lg(CFU/g) and 1.13 lg(CFU/g) respectively, and PPO activity decreased from 11.40 U to 6.31 U. The dough quality properties, such as stability, extensibility, and starch viscosity, improved significantly under different microwave conditions. Confocal laser scanning microscopy(CLSM) images indicated that starch and proteins aggregated gradually in treated flour, altering rheological properties of dough. From the results of scanning electron microscopy(SEM), microwave treatment led to the appearance of disrupted structure in the gluten proteins, but the secondary structure of proteins altered slightly. Rheological properties of dough confirmed that the microwave treatment greatly affected processing characteristics of wheat flour products, with significant advantageous consequences on product quality, especially for textural properties of FWN. Furthermore, FWN darkening could be inhibited noticeably after microwave treatment, thereby prolonging its shelf life. Therefore, microwave treatment could thus be an effective, practical technology to produce low-bacterial flour and thereby enhance its product quality.

Prediction of Protein OmpH in Structure of C47-8 Pasteurella multocida

[Objective] This study aimed to predict the structure of protein OmpH from Pasteurella multocida C47-8 （PmC47-8） strain of yak. [Method] Online BLAST, signal peptide prediction, secondary structure prediction and protein characteristics of sequencing result of gene OmpH from PmC47-8 strain were analyzed. [Result] The similarities of gene OmpH from PmC47-8 with the published 81 OmpH genes were between 84% and 99%; a signal peptide was found with the cleavage sites between 20 and 21 in the polypeptide; secondary structure prediction showed that folding structure accounted for 49.8% and loop structure for 50.2%; it predicted that there were 7 O-glycosylation sites in OmpH protein with the amino acid residual sites of 2, 45, 48, 330, 716, 721, 723, respectively, and 2 N-glycosylation sites with the amino acid residual sites of 15 and 35. [Conclusion] This study lays the foundation for the study on the immunity of OmpH gene from yak.

A GASVM Algorithm for Predicting Protein Structure Classes

The research methods of protein structure prediction mainly focus on finding effective features of protein sequences and developing suitable machine learning algorithms. But few people consider the importance of weights of features in classification. We propose the GASVM algorithm (classification accuracy of support vector machine is regarded as the fitness value of genetic algorithm) to optimize the coefficients of these 16 features (5 features are proposed first time) in the classification, and further develop a new feature vector. Finally, based on the new feature vector, this paper uses support vector machine and 10-fold cross-validation to classify the protein structure of 3 low similarity datasets (25PDB, 1189, FC699). Experimental results show that the overall classification accuracy of the new method is better than other methods.

FTIR Analysis of Protein Secondary Structure in Cheddar Cheese during Ripening

Proteolysis is one of the most important biochemical reactions during cheese ripening.Studies on the secondary structure of proteins during ripening would be helpful for characterizing protein changes for assessing cheese quality.Fourier transform infrared spectroscopy(FTIR),with self-deconvolution,second derivative analysis and band curve-fitting,was used to characterize the secondary structure of proteins in Cheddar cheese during ripening.The spectra of the amide I region showed great similarity,while the relative contents of the secondary structures underwent a series of changes.As ripening progressed,the α-helix content decreased and the β-sheet content increased.This structural shift was attributed to the strengthening of hydrogen bonds that resulted from hydrolysis of caseins.In summary,FTIR could provide the basis for rapid characterization of cheese that is undergoing ripening.

Does mRNA structure contain genetic information for regulating co-translational protein folding？

Currently many facets of genetic information are illdefined. In particular, how protein folding is genetically regulated has been a long-standing issue for genetics and protein biology. And a generic mechanistic model with supports of genomic data is still lacking. Recent technological advances have enabled much needed genome-wide experiments. While putting the effect of codon optimality on debate, these studies have supplied mounting evidence suggesting a role of m RNA structure in the regulation of protein folding by modulating translational elongation rate. In conjunctions with previous theories, this mechanistic model of protein folding guided by m RNA structure shall expand our understandings of genetic information and offer new insights into various biomedical puzzles.

Characterization of Protein in Old Myocardial Infarction by FTIR Micro-spectroscopy

The aim of the present study was to assess whether Fourier transform infrared spectrometry (FTIR) micro-spectroscopy could produce distinct spectral information on protein of old myocardial infarction (OMI) and to set them as molecular markers to diagnose atypical OMI. Paraffin-embedded heart samples were derived from victims dying of OMI. In combination with histological stain, FTIR and infrared micro-spectroscopy, the characteristics of OMI were analyzed morphologically and molecularly. The most relevant bands identified were the amide A, B, Ⅰ and Ⅱ, showing crucial spectral differences between apparent normal region and OMI region, including the peak position blue shift and the increased intensity of OMI, moreover relative increase in α-helix and decrease in β-sheet of protein secondary structures in OMI. Comparing to single spectral band, the I1650/I1550 ratio was increased and rationally used as a molecular marker for diagnosing OMI. These novel preliminary findings supported further exploration of FTIR molecular profiling in clinical or forensic study, and were in accordance with histopathology.

Proteins:From sequence to structure

Protein sequences as special heterogeneous sequences are rare in the amino acid sequence space. The specific sequen- tial order of amino acids of a protein is essential to its 3D structure. On the whole, the correlation between sequence and structure of a protein is not so strong. How well would a protein sequence contain its structural information？ How does a sequence determine its native structure？ Keeping the globular proteins in mind, we discuss several problems from sequence to structure.

Construction of eukaryotic expression plasmid containing HCV NS3 segment and protein expression in human HL-7702 hepatocytes

AIM： To construct the eukaryotic expression plasmid containing HCV NS3 segment and to analyze the expression of NS3 protein in normal human hepatocyte HL-7702.METHODS： We amplified HCV NS3 fragment from plasmid pBRTM/HCV 1-3011 containing the whole length of HCV genome, recombined it with expression vector pcDNA3.1（-） to form the eukaryotic expression vector pcDNA3.1（-）/NS3, and transfected human HL-7702 hepatocytes with the recombined plasmid by cationic polymers. The expressed HCV NS3 protein was detected and analyzed by immunohistochemical method and Western blot.RESULTS： The amplified NS3 fragments had correct molecule weight and sequence. The successfully constructed eukaryotic expression plasmids were transfected to HL-7702 cells. The expressed NS3 proteins had correct molecular weight 70000.CONCLUSION： Eukaryotic expression vector pcDNA3.1 （-）/NS3 containing NS3 segment of HCV can be constructed, the sequence of NS3 fragments is consistent with the template. Normal human HL-7702 hepatocytes can efficiently express specific HCV NS3 protein in vitro.

HMM in Predicting Protein Secondary Structure

We introduced a new method---duration Hidden Markov Model (dHMM) to predicate the secondary structure of Protein. In our study, we divide the basic second structure of protein into three parts: H (a-Helix), E (B-sheet) and O (others, include coil and turn). HMM is a kind of probabilistic model which more thinking of the interaction between adjacent amino acids (these interaction were represented by transmit probability), and we use genetic algorithm to determine the model parameters. After improving on the model and fixed on the parameters of the model, we write a program HMMPS. Our example shows that HMM is a nice method for protein secondary structure prediction.