Preparation of human monoclonal antibody against SARS-CoV-2 spike protein using single B cell / 中国生物制品学杂志

@#Abstract：Objective To prepare human monoclonal antibody against spike protein（S protein）of severe acute respiratory syndrome coronavirus 2（SARS⁃CoV⁃2）by using single B cell，and determine its neutralizing activity. Methods Venous blood with high antibody level was collected from people immunized with inactivated SARS⁃CoV⁃2 vaccine（Vero cells） twice，of which peripheral blood mononuclear cells（PBMCs）were isolated by lymphocyte stratified fluid and used to isolate single B cell expressing S protein antibody by magnetic beads coupled with S1 protein. Variable region genes of IgG heavy chain and light chain were amplified by nested PCR after reverse transcription of single B cell，which were connected with CMV promoter，IgG leader sequence，IgG constant region and polyA sequence by overlapping PCR to construct antibody linear expression cassette. Linear expression cassette of the heavy chain and light chain from the same B cell was transfected to HEK293T cells to express human monoclonal antibody of SARS⁃CoV⁃2 S protein. Immunoreactivity was detected by immuno⁃ fluorescence while neutralizing activity by pseudovirus neutralization test. Results A total of 26 monoclonal antibodies against SARS⁃CoV⁃2 S protein were expressed，which showed heavy chain and light chain protein bands of IgG antibody at

Construction and identification of recombinant adenovirus expressing S protein receptor binding domain and N protein of SARS-CoV-2 Delta variant / 中国生物制品学杂志

@#ObjectiveTo construct and identify a recombinant adenovirus expressing S protein receptor binding domain(RBD)and N protein of severe acute respiratory symptom coronavirus 2(SARS-CoV-2)Delta variant.MethodsThe RBD and N gene fragments of SARS-CoV-2 were cloned into pcDNA3.0BA vector respectively to construct recombinant plasmid pcDNA3.0BA-RBD-N. The RBD-CMV-N fragment was amplified by PCR and inserted into shuttle vector pShuttle-CMV. The shuttle plasmid pShuttle-RBD-N was then homologously recombined with pAdeasy-1 to obtain recombinant plasmid pAdeasy-1-RBD-N,which was transfected into HEK293 cells for recombinant adenovirus Ad-RBD-N packaging. The transcription of RBD and N genes of recombinant adenovirus in HEK293 cells was detected by RT-PCR,while the expre-ssion of RBD and N proteins by Western blot and immunofluorescence assay. 12 female BALB/c mice were immunized with Ad-RBD-N by intramuscular injection at a dose of 5 × 109copies per mouse. Blood samples were collected 14 d after immunization,and the serum antibody titers were measured by ELISA.ResultsThe RBD and N genes of recombinant adenovirus were transcribed normally in HEK293 cells,and the RBD and N proteins were expressed normally in MA104 cells. Mice immunized with the recombinant adenovirus produced specific IgG antibodies against RBD and N proteins.ConclusionThe recombinant adenovirus expressing S protein RBD and N protein of SARS-CoV-2 Delta variant was succe-ssfully constructed,which laid a foundation of the follow-up research on Delta variant vaccines.

Epidemiological and molecular characteristics of SARS-CoV-2 Delta variant-caused pneumonia in Henan Province in 2021 / 中华微生物学和免疫学杂志

Objective:To analyze and summarize the epidemiological and molecular characteristics of SARS-CoV-2 Delta variant, a variant of concern (VOC), in Henan Province in 2021 in order to provide a basis for epidemic prevention and control.Methods:According to the feedback of sequencing results from Chinese Center for Disease Control and Prevention, 111 patients infected with SARS-CoV-2 Delta VOC were selected from the Henan imported and local cases in 2021. Basic patient information was obtained from the pandemic website. The differences in age, gender, vaccination history, the number of vaccine doses and different clinical types were analyzed. Moreover, the differences in RT-qPCR results of ORF1 ab gene and N gene Ct values between cases of different genders and symptoms were analyzed statistically. Sequencing results of the nucleotide and S protein mutation sites were analyzed. Results:There was no significant difference in the gender distribution of 111 cases between different age groups (χ 2=2.217, P=0.529). There was also no significant difference in clinical types between patients with different vaccination history (χ 2=12.074, P=0.209). The Ct values of most SARS-CoV-2 nucleic acid-positive specimens were distributed in the lower range and the viral loads were higher. The difference in the Ct value of ORF1 ab gene between different gender groups was not statistically significant (χ 2=1.646, P=0.439), but were significantly different among asymptomatic, mild, normal, and severe cases (χ 2=13.257, P=0.039). There was no significant difference in N gene Ct value among cases of different genders or different symptoms (all P>0.05). The 111 patients in this study were mainly found through close-contact screening and full-staff nucleic acid screening and accounted for 62.2% (69 cases) of the total. The sequencing length coverage was basically greater than 99% (accounting for 90.1%, 100/111); the total number of nucleotide mutation sites was mostly in the range of 46-50 (86.4%, 89/103); the total number of S protein mutation sites was mostly 12 (82.5%, 85/103). The 103 Delta mutants all contained nine mutation sites, which were T19R, R158G, L452R, T478K, D614G, P681R, D950N, E156del and F157del, with a mutation rate of 100%. Conclusions:People were highly susceptible to the SARS-CoV-2 Delta in Henan Province in 2021. High viral load and increase in the ORF1 ab gene load would aggravate the clinical symptoms.

Novel epitope based peptides for vaccine against SARS-CoV-2 virus: immunoinformatics with docking approach

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative viral strain for the contagious pandemic respiratory illness in humans which is a public health emergency of international concern. There is a desperate need for vaccines and antiviral strategies to combat the rapid spread of SARS-CoV-2 infection.Methods: The present study based on computational methods has identified novel conserved cytotoxic T-lymphocyte epitopes as well as linear and discontinuous B-cell epitopes on the SARS-CoV-2 spike (S) protein. The predicted MHC class I and class II binding peptides were further checked for their antigenic scores, allergenicity, toxicity, digesting enzymes and mutation.Results: A total of fourteen linear B-cell epitopes where GQSKRVDFC displayed the highest antigenicity-score and sixteen highly antigenic 100% conserved T-cell epitopes including the most potential vaccine candidates MHC class-I peptide KIADYNYKL and MHC class-II peptide VVFLHVTYV were identified. Furthermore, the potential peptide QGFSALEPL with high antigenicity score attached to larger number of human leukocyte antigen alleles. Docking analyses of the allele HLA-B*5201 predicted to be immunogenic to several of the selected epitopes revealed that the peptides engaged in strong binding with the HLA-B*5201 allele.Conclusions: Collectively, this research provides novel candidates for epitope-based peptide vaccine design against SARS-CoV-2 infection.

Overview of novel coronavirus infection and replication / 生物工程学报

Coronaviruses are a type of positive-sense single-stranded RNA virus with envelope and widely exist in nature to cause respiratory infectious diseases. The novel coronavirus is a new outbreak virus that is susceptible to all people. Up to now, the disease has been widely spread in the world and poses a great threat to public health. In this review, the genomic features, key proteins, host infection and replication of coronaviruses and novel coronaviruses are reviewed in order to provide theoretical basis for the study of the pathogenic mechanism of virus infection on host cells and to provide basic support for the development of specific antiviral drugs.

Screening novel inhibitors targeting SARS-CoV-2 S protein-ACE2 interaction based on molecular docking / 中草药

Objective: To screen inhibitors targeting SARS-CoV-2 S protein-ACE2 interaction by molecular docking. Methods: Candidate natural products were collected from Selleck China natural product library (Catalog No. L1400, 2 054 natural products). The structure of SARS-CoV-2 S protein-ACE2 had been determined by Qiang Zhou team (PDB: 6M17). The molecular docking was performed by Discovery Studio. Results: Based on the virtual amino acid mutation experiment which determined the key amino acids, the binding cavity was created. Then, 11 compounds were screened out from the natural compound library: digitonin, Lonicera grisea saponin A, forsythiaside B, L. grisea saponin B, Dipsacus asperges saponin B, hederacoside D, platycodon D, echinacoside, ginsenoside Rb2, ginsenoside Rc, and chlorogenic acid C. Conclusion: The 11 potential inhibitors targeting SARS-CoV-2 S protein-ACE2 interaction were screened out from natural products library, which provides a reference for the research of new anti SARS-CoV-2 drugs.

Sequence analysis of the HBV S protein in patients with chronic hepatitis B / 中华检验医学杂志

Objective To analysis the frequency and mutational patterns of HBV(hepatitis B virus) S protein in the different phases of chronic hepatitis B patients, exploring the clinical implication of these mutations in CHB. Methods The difference of S protein mutation rates and mutation site among each groups were analyzed by cases-comparison. 112 cases of chronic hepatitis B patients in Shanghai Shu-guang hospital were enrolled in this study; they were divided into four groups, immune-tolerance group [IT, 36 cases, male 20 cases,female 16 cases,age 28.00 (25.00,30.75)years], immune-clearance group [IC, 28 cases, male 17 cases,female 11 cases,age 29.00 (25.25,31.75) years], low-replication group [LR, 25 cases, male 17 cases,female 8 cases,age 39.00 (35.00,45.50) years], reactivation group [RE, 23 cases, male 15 cases,female 8 cases, age 43.00 (36.00, 48.00) years]. DNA sequencing was used to detection HBV S protein gene mutation. Difference between categorical variables were analyzed using Chi-square test, Non-parametric test (kruskal-wallis H and Mann-Whitney U) was used for non-normal distribution data. Results Naturally occurring MHR (major hydrophilic region) variants was 16.67％ (6/36 cases) in IT group, preferentially within"a"determinant in the first loop, compared with IC (21.43％,6/28 cases), LR (20.00％,5/25 cases) and RE (34.78％,8/23 cases), there was no statistics Significance (χ2=2.814, P=0.421), while variant frequencies outside the MHR were 11.11％ (4/36 cases) and 14.29％ (4/28 cases) in IT and IC group, viral diversity gradually increased during LR phase with 24.00％(6/25 cases) of mutation frequency, and peak at RE phase (52.17％, 12/23 cases) with a statistics significance (χ2=15.041, P=0.002). Conclusion Amino acid substitutions is not only the consequence of the pressure of host immunity, it may be a probably cause of disease reactivation.

Advances in reverse genetics to treat porcine epidemic diarrhea virus / 生物工程学报

Porcine epidemic diarrhea virus (PEDV) is one of the major etiologies responsible for the acute, highly contagious disease in the digestive tract of pigs, especially neonatal piglets. Since PEDV was first identified in Europe in the late 1970s, it has resulted in significant economic losses in many Asian swine-raising countries, including China. Recently, reverse genetics techniques including targeted RNA recombination, bacteria artificial chromosome system and in vitro ligation have been successfully used to manipulate the genome of PEDV, which providing new strategies for the clear delineation of the functions of the viral proteins, the mechanisms behind PEDV pathogenesis and the design of novel vaccines against PEDV. Here, we review the progresses of different reverse genetics platforms developed for PEDV and their applications, covering the roles of trypsin in PEDV propagation, functions of S and ORF3 protein and the development of next generation PED vaccines, and the perspectives of reverse genetics for PEDV.

Epidemiological and molecular virological characteristics of HBV infected patients of hospital with simultaneously positive HBsAg and anti-HBs / 临床检验杂志

Objective To investigate the epidemiological and molecular virological characteristics in HBV-infected patients with copositive HBsAg and anti-HBs.Methods HBV serological markers were analyzed in 52 070 specimens.The epidemiological characteristics of HBsAg and anti-HBs simultaneously positive patients (the experimental group) and HBsAg positive and auti-HBs negative patients (the control group) were compared.The S protein of HBV coding region was amplified by semi-nested PCR and sequenced.The statistical differences between the two groups were compared in different gene regions,genotypes and different clinical diagnosis.Results HBsAg was positive in 20.40％ (10 621/52 070) of all specimens.In the patients with positive HBsAg,2.48％ (263/10 621) was positive anti-HBs.The prevalence of co-positive HBsAg and auti-HBs was higher in aged 0 to 9 years and greater than or equal to 80 years than that in other age,and the prevalence of positive HBsAg and negative anti-HBs was completely opposite.The mutation rate of S protein in the experimental group was significantly higher than that in the control group (1.52％ vs 0.81％,P ＜0.01) with the mutation in the major hydrophilic region (MHR) (1.68％ vs 0.57％,P ＜0.01).The mutation rates of S protein of HBV carriers,chronic hepatitis B (CHB) patients and patients with liver cirrhosis (LC) in the experimental group were significantly higher than those in the control group (1.47％ vs 0.65％,1.28％ vs 0.84％,2.21％ vs 0.44％,P ＜0.05,respectively),except for the patients with hepatocellular carcinoma (HCC) (1.97％ vs 2.21％,P ＞ 0.05).Conclusion Co-positive HBsAg and anti-HBs in HBV-infected patients was more common in HBsAg positive patients aged 0 to 9 years and greater than or equal to 80 years than the others.Coexistence of HBsAg and anti-HBs in HBV-infected patients may relate to immune escape caused by mutation of S protein (mainly MHR).The mutation rates of S protein in the two groups of patients,co-positive HBsAg and anti-HBs and the positive HBsAg combined with negative anti-HBs,were associated with the stage of liver disease.

Agammaglobulinemia ligada al x o de bruton / X-linked agammaglobulinemia or bruton´s disease

La agammaglobulinemia ligada al X (ALX) o de Bruton es una inmunodeficiencia primaria que generalmente se manifiesta en los primeros meses de la vida, cuando disminuyen las concentraciones séricas de las inmunoglobulinas maternas. Se caracteriza por infecciones recurrentes y ausencia total o niveles muy bajos de inmunoglobulinas. Se reporta el caso de un niño de 5 años de edad con historia de procesos infecciosos severos recurrentes de comienzo a los 18 meses de nacido: shigellosis, infecciones respiratorias bacterianas, bronconeumonías, conjuntivitis, sinusitis, meningoencefalitis en tres ocasiones (dos de etiología viral y una de etiología bacteriana), otitis media supurativa crónica, giardiasis de evolución tórpida y lesiones sépticas en piel por pseudomona aeruginosa y estafilococo dorado. Durante el curso de los procesos infecciosos se diagnosticó una enfermedad autoinmune (psoriasis). El estudio inmunológico realizado mostró niveles extremadamente reducidos de las inmunoglobulinas séricas: IgG 0,00 mg/L (370 - 1 400 mg/L); IgA 0,08 g/L (50 - 230 mg/L); e IgM 0,07 g/L (30 - 170 mg/L), así como células B CD19+ en sangre periférica casi ausentes, con un valor de 0,12 por ciento (VN: 21 - 44 por ciento ). Se estableció el diagnóstico de agammaglobulinemia ligada al X o de Bruton. El paciente recibió tratamiento con inmunoglobulina humana por vía endovenosa con mejoría clínica evidente...

X-linked agammaglobulinemia (XLA) or Bruton disease is a primary immunodeficiency, which typically appears in the first months of life, when serum concentrations of maternal immunoglobulins decrease. It is characterized by recurrent infections and total absence or very low levels immunoglobulin. We report a 5-year-old boy with a history of recurrent severe infectious processes beginning at 18 months of age: shigellosis, bacterial respiratory infections, bronchopneumonia, conjunctivitis, sinusitis, meningoencephalitis three times (two of viral etiology and one of bacterial etiology), chronic suppurative otitis media, giardiasis with torpid evolution and septic skin lesions caused by Pseudomona aeruginosa and Staphylococcus aureus. During the course of infectious processes an autoimmune disease (psoriasis) was diagnosed. Immunological study showed extremely low levels of serum immunoglobulins: IgG 0.00 mg / L (370 - 1 400 mg / L), IgA 0.08 g / L (50 - 230 mg / L), and IgM 0, 07 g / L (30 - 170 mg / L) and CD19 + B cells in peripheral blood almost absent, with a value of 0.12 percent (VN: 21 - 44 percent). Diagnosis of X-linked agammaglobulinemia or Bruton disease was established. The patient was treated with intravenous human immunoglobulin with obvious clinical improvement...

The Usefulness of P504S/34betaE12 Immunostaining for the Detection of Prostate Cancer / 대한비뇨기과학회지

PURPOSE: We evaluated the significance of the P504S expression in prostate cancer and also the usefulness of the P504S/34betaE12 combined immunostaining method for diagnosing prostate cancer, and we did this by performing histological analysis of needle biopsy specimens. MATERIALS AND METHODS: Prostate tissue specimens were obtained from 83 patients with clinically suspected prostate cancer. A total of 54 prostate needle biopsy specimens were immunostained with an enzyme commonly overexpressed in prostate cancer(P504S) and also with an antibody against a basal cell marker(34betaE12). A total of 83 cases were immunostained with 34betaE12, including 29 cases that were stained with only with HMW- CK(34betaE12). RESULTS: P504S immunostaining was positive in 96.3%(26 of 27 cases) of the prostate cancer specimens. 34betaE12 immunostaining was positive in 97.2%(35 of 36 cases) of the benign prostate tissues. Of the 30 P504S positive immunostaining cases, 26 cases were prostate cancers, 3 cases were ASAP and 1 case was ASAP+PIN. Of the 36 34betaE12 positive immunostained cases, 35 cases were benign and 1 case was ASAP. In the P504S(+)/34betaE12(-) cases, there are no benign prostate lesions. There are no benign prostate lesions in the P504S(-)/34betaE12(+) cases, and all the cases were benign. There were no statistical correlations between the grade of P504S staining and the clinical parameters such as serum PSA, the clinical stage and the Gleason scores. CONCLUSIONS: Combining P504S as a positive marker for prostate cancer with 34betaE12 as a negative marker might improve the diagnostic performance.

The Functional Motif of SARS-CoV S Protein Involved in the Interaction with ACE2 / 中国病毒学

SARS-CoV is a newly discovery pathogen causing severe acute respiratory problems.It has been established that the S protein in this pathogen plays an important rule in the adsorption and penetration of SARS-CoV into the host cell by interaction with the ACE2 receptor.To determinant which functional motif of the S protein was involved in the interaction with ACE2,seven truncated S proteins deleted from the N or C terminal were obtained by an E.coli expression system and purified by column chromatography to homogeneity.Each truncated S protein was fixed on to the well of an ELISA plate and an interaction was initiated with the ACE2 protein.The adsorption were quantified by ELISA,and the results indicated that amino acids from 388 to 496 of the S protein was responsible for the interaction with the ACE2 receptor,and the interaction could be completely disrupted by an antibody specific to these amino acids.Deletions adjacent to this domain did not appear to have a significant impact on the interaction with ACE2,suggesting that the S protein of SARS-CoV could be developed as a vaccine to prevent the spread of SARS-CoV.

Expression of angiotensin-converting enzyme 2 in rat pancreas / 基础医学与临床

Objective To investigate the expression of angiotensin-converting enzyme 2(ACE2) in rat endocrine and exocrine tissue of pancreas and to compare its expression level with that in other visceral tissues. Methods The expression of ACE2 in rat visceral organs was detected by immunohistochemistry and reverse transcriptase polymerase chain reaction (RT-PCR). Results ACE2 was positively expressed in all the test tissues, including pancreas, heart, liver, kidney, lung, spleen, duodenum, ileum and bladder. The expression of ACE2 mRNA was detected in whole pancreas and ACE2 protein was found in both endocrine tissue and exocrine tissue of rat pancreas, while in the endocrine tissue much less ACE2 protein was expressed. The absorbance value of exocrine tissue of rat pancreas increased significantly as compared to that in pancreatic islet. Conclusion Our data demonstrated a tissue specific pattern of ACE2 expression in rat pancreas and this indicated ACE2 might be present in islet of pancreas of human, which might cause the elevation of blood glucose levels in SARS patients. Since ACE2 works as a negative regulator of the angiotensin system, the expression of ACE2 protein in endocrine tissue of pancreas may facilitate the treatment of diabetes.

The possible mechanism of the acute lung injury induced by S protein of SARS-CoV / 医学研究生学报

Objective: To investigate the role of the recombinant S protein of SARS-CoV in the induction of chemokine IP-10 and other cytokines in airway epithelial cells and immunocytes. Methods: Using insect-baculovirus expression system and Nickel affinity Magnet Beads, S protein of SARS-CoV was produced and then used to stimulate cultured human bronchial epithelial cells (16HBE), human peripheral blood mononuclear cells(PBMC), human peripheral blood monocytes and alveolar macrophages. The levels of IP-10 and the cytokines involved in immunoreaction in response to virus infection were detected in the supernatants of those cells cultured with the S protein by liquid chip system. Results: Under normal condition, no detectable IP-10 was found in 16HBE. A high level of IP-10(79.97? 13.81) pg/ml was detected in the 16HBE 12 hrs after being treated with the S protein, and the induction of IP-10 by S protein displayed at a significant quantity-effect reaction, but not in PBMC, monocytes and alveolar macrophages. In contrast, IFN-? was able to induce the production of IP-10 in either 16HBE or the immunocytes. Conclusion: 1.S protein of SARS-CoV can induce a high level of IP-10 in lung epithelial cells at early stage after the virus infection, which may initiate the process of the immune damage in the lung. 2. S protein of SARS-CoV induces the production of IP-10 by a way of IFN-? independent.

Effect of alkaline pH on sunflower 11S protein.

The effect of alkaline pH on sunflower 11S protein has been monitored by the techniques of ultracentrifugation, polyacrylamide gel electrophoresis, turbidity, viscosity, ultraviolet absorption spectra and fluorescence spectra· Both ultracentrifugation and polyacrylamide gel electrophoresis show the dissociation of the protein with increase in pH. Turbidity values decrease with pH while viscosity increases· With increase in pH absorbance of the protein solution increases and there is a red shift in the absorption maximum. Fluorescence quenching and a red shift in the emission maximum are also observed. Both dissociation and denaturation of the protein occur. Analysis of turbidity, viscosity and fluorescence data suggests that apparently denaturation follows dissociation.

SARS-CoV S protein induces chemokine IP-10 expression in airway epithelial cells / 中国病理生理杂志

AIM: To investigate the molecular mechanism by which the SARS-CoV S protein induces chemokine IP-10 in airway epithelial cells.METHODS: cDNA microarrays were used to screen the gene expression profiles of human bronchial epithelial cells(16HBEs) stimulated by SARS-CoV S protein.In addition,RT-PCR,EMSA,and Western blotting were performed to analyze the phosphorylation of JAK/STAT signal pathway.The changes of IRF-1 and IP10 gene expression and the influence by the corresponding inhibitors were analyzed.RESULTS: IRF-1,a key transcription factor of the JAK/STAT signal pathway,was activated in human bronchial epithelial cells after stimulation by the S protein of SARS-CoV.The IP-10 gene expression was detected 2 h following the phosphorylation of STAT1 after 15 min,which was blocked by STAT1or JAK2 inhibitors.Electrophoretic mobility shift assay(EMSA) demonstrated that the nuclear proteins bound to ISRE and GAS but not NF-?B DNA motif.CONCLUSION: The SARS-CoV S protein induces IP-10 gene expression in human bronchial epithelial cells through activation of the JAK/STAT signal pathway,suggesting that the JAK/STAT signal pathway activated by virus plays key roles in virus infection related acute lung injury.

Prediction of the B cell epitope for the S protein of SARS coronavirus / 第三军医大学学报

Objective To predict the B cell epitopes for S protein of severe acute respiratory syndrome (SARS) coronavirus. Methods Based on SARS coronavirus genome sequence and the analysis of the flexible regions of secondary structure for S protein, the B cell epitopes for S protein were predicted by methods of Kyte Doolittle hydrophilicity plot, Emini, and Jameson Wolf. Results The computer predicted most possible epitopes for S protein were located within or nearby its N terminal No. 91-98, 1121-1153 and 185-193. The N terminal No. 1050-1059, 752-765, 41-50, 666-674, 264-287, 340-348, 408-416, and 424-439 may be the possible B cell epitopes. Conclusion Prediction of the B cell epitopes for the S protein based on multiple parameters is helpful for the identification of B cell epitopes using experimental methods.

Effect of denaturants on the 11S protein of guar seed.

The effect of sodium dodecyl sulphate, urea or guanidinium hydrochloride on the sedimentation velocity, viscosity, ultra violet spectra and fluorescence spectra of the 11S protein of guar seed has been determined. Sodium dodecyl sulphate dissociates the protein directly to the 2S protein, whereas urea or guanidinium hydrochloride produces an intermediate 7S protein. These reagents denature the protein also. Both the dissociative and the denaturation effect follow the order, sodium dodecyl sulphate > guanidinium hydrochloride > urea when the concentration are expressed as mols per litre. The denatured states in the three cases probably differ.

Isolation and characterization of the 7S protein from glanded cottonseed (Gossypium herbacium).

The major protein from glanded cottonseed has been isolated in a homogeneous form. Its S20, w value at 1% protein concentration is 6S in 1 Μ NaCl solution. It contains 1% carbohydrate and is free from phosphorus, gossypol (bound or free) and nucleic acid impurities. It consists of atleast seven non-identical subunits. The protein has an ultraviolet absorption maximum at 278 nm and fluorescence excitation and emission maxima at 280 nm and 325 nm respectively. Optical rotatory dispersion and circular dichroism measurements indicate that the protein consists predominantly of β-structure and random coil. The observed near-ultraviolet circular dichroic bands can be attributed to tyrosine, phenylalanine and tryptophan residues of the protein.

Effect of denaturants on the oligomeric structure of the 11 S protein of sunflower (Helianthus annum).

The effect of sodium dodecyl sulphate, urea, guanidinium hydrochloride and heat on the oligomeric structure of the 11 S protein of sunflower has been determined. Sodium dodecyl sulphate directly dissociates the protein to 2 S subunits, whereas urea and guanidinium hydrochloride dissociate it through an intermediate 7 S protein. Heating the protein at 90°C for 20 min caused dissociation of the 11 S protein, without any precipitation.