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Ren H.-W.,National Dairy Engineering & Technical Research Center | Ren H.-W.,Northeast Agricultural University | Zhang W.-S.,National Dairy Engineering & Technical Research Center | Zhang W.-S.,Northeast Agricultural University | And 4 more authors.
Guang Pu Xue Yu Guang Pu Fen Xi/Spectroscopy and Spectral Analysis | Year: 2015

To obtain a structural basis for the β-casein in Chinese human milk, structural transitions of the β-casein in response to variation of pH were investigated using Raman and circular dichroism (CD) spectroscopy. Both methods indicated that the secondary structures of β-casein in the solution were induced by the pH. Secondary structural analysis of β-casein by CD spectroscopy yielded 0.5%~2% α-helical, 16%~18% β-sheet, 30%~34% β-turn and 49%~51% random coil contents. Another result was that as pH increases, these structures change. Several distinct transitions were observed by circular dichroism in α-helix at pH 8 and pH 10. Raman spectrum also showed random coil as the major secondary structure in native β-casein, for the characteristic band of the β-casein amide I was at 1662 cm-1. Calculations from I850/I830 suggested that the tyrosine residues of β-casein tended to “exposure”. CD and Raman spectra both showed that at neutral and alkaline pH the β-casein existed predominantly in random coil conformation, and the proportion of α-helix was higher at pH 8 than under other pH conditions. Over the range of pH studied, the sheet and turn areas remained relatively constant, and in the condition of pH 8, the content of α-helical was higher than in the other pH conditions. ©, 2014, Science Press. All right reserved.

Ren H.,National Dairy Engineering & Technical Research Center | Ren H.,Northeast Agricultural University | Li C.,Northeast Agricultural University | Liu N.,National Dairy Engineering & Technical Research Center | And 3 more authors.
Journal of Chinese Institute of Food Science and Technology | Year: 2015

It has been showed there are 12 genetic variants of β-casein in Bos taurus, and this kind of phenomenon might inherent in Homo sapiens. To confirm whether there was diversity between the primary structure of Chinese human β-casein and the data of NCBI database, and to verify the amino acid sequence of Chinese human β-casein that we reported previously. The total RNA extracted from normal breast tissue of a Chinese female was processed by RT-PCR to produce the human β-casein Coding Sequences region. The PCR product was cloned into vector T for sequence analysis, this sequence was submitted to GeneBank database (accession number: KF 751868). BLAST result of this sequence and the human β-casein CDS region published in NCBI database shows that the homology was 100%, such highly homology between different races suggests that this gene is conserved. The bioinformatic analyses and forecasts of the nucleotide sequence and the deduced amino acid sequence were conducted: the theoretical pI was 5.52 and the protein contained no span membrane structures, and there existed high hydrophobicity region in the amino acid sequence. The recombinant plasmid was cloned into the vector PET-30a-GST, induced by IPTG at 37 degrees Celsius for 4h, the fusion protein expressed in Escherichia coli (E. coli) BL21 (DE3) mainly existing in the inclusion bodies. The recombinant human β-casein was purified using NI-NTA affinity chromatography, the result of SDS-PAGE indicated that the prokaryotic expression vector PET-30a-GST-CSN2 and the human recombinant CSN2 were gained. The whole work laid a solid foundation for the study of Chinese human β-casein. ©, 2015, Chinese Institute of Food Science and Technology. All right reserved.

Huang Y.,National Dairy Engineering & Technical Research Center | Ren H.,National Dairy Engineering & Technical Research Center | Liu B.,National Dairy Engineering & Technical Research Center | Liu N.,National Dairy Engineering & Technical Research Center | And 2 more authors.
Se pu = Chinese journal of chromatography / Zhongguo hua xue hui | Year: 2013

The selective precipitation of whole casein from skimmed milk was achieved by the addition of calcium salt under acidic pH. The effects of pH, centrifugal force and final concentration of CaCl2 on the separation of casein were studied by measuring the purity of final products using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The results showed that casein with the highest purity could be obtained with the pH of 4.3, the centrifugal force of 10 400 g and the final concentration of CaCl2 of 60 mmol/L. The casein was processed with DEAE anion exchange chromatography and three peaks were obtained. Then the third peak (peak III) was identified with Western-Blot method and matrix-assisted laser desorption/ionization time of flight/time of flight mass spectrometry (MALDI-TOF/TOF MS). The identification of Western-Blot showed that peak III can combine with the specificity of human milk beta-casein antibody, and it is proved to be human milk beta-casein. The fingerprints of peak III were nalyzed by Mascot searching, and the sequence coverage was 50%, further supporting it is human milk beta-casein. In conclusion, an effective method to obtain human milk beta-casein from milk samples through DEAE anion exchange chromatography was established, and it is suitable for the proteomics research requirements of the beta-casein from human milk.

He Y.,Northeast Agricultural University | He Y.,National Dairy Engineering & Technical Research Center | Ren H.,Northeast Agricultural University | Ren H.,National Dairy Engineering & Technical Research Center | And 8 more authors.
CYTA - Journal of Food | Year: 2016

The fatty acid (FA) composition in human milk, bovine milk and infant formula from the northeast region of China was analyzed by gas chromatography (GC). The content of linoleic acid (LA, C18:2n6) in bovine milk (2.53 %) was lower than human milk (25.58 %) and formula milk (20.56 %). The trends of LA and α-linolenic acid (ALA, C18:3n3) in human milk increased significantly throughout the lactation while a decreasing trend for LA was observed and the difference of ALA was not significant in both bovine and formula milks. The proportion of docosahexaenoic acid (DHA, C22:6n3) and arachidonic acid (ARA, 20:4n6) in human milk and formula were declining with the change of lactation or infant age. We suggest there should be more detailed distinction with reference to human milk in the infant formula in order to meet infant’s personalized nutrition, especially the FA composition in the 1st month after delivery. © 2016 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group

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