Characterization of the bovine milk proteome in early-lactation Holstein and Jersey breeds of dairy cows

Tacoma, R.; Fields, Julia; Ebenstein, David B.; Lam, Ying‐Wai; Greenwood, S.L.

doi:10.1016/j.jprot.2015.09.024

Cited by 68 publications

(76 citation statements)

References 57 publications

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“…However, the use of mammary glands as bioreactors has some disadvantages: (1) only TG female animals can produce heterologous proteins from their milk; (2) TG female animals can synthesize foreign proteins in their milk only when they are at the lactation stage; (3) in some animal species the lactation period is short and hence merely a small amount of transgene-encoded proteins can be produced from milk; (4) in addition, milk usually contains a large amount of diverse endogenous proteins, which complicates the isolation of high-purity foreign proteins. For example, human milk contains more than 1,600 proteins and has a total protein concentration of 13 mg/ml5, while cattle milk carries approximately 1,000 proteins and has 30 mg/ml of total proteins6. Hence, there is a need to develop new bioreactor systems for the efficient production of valuable proteins and here we report salivary glands as an excellent alternative.…”

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confidence: 73%

Production of functional human nerve growth factor from the saliva of transgenic mice by using salivary glands as bioreactors

Zeng

Zhu

et al. 2017

Sci Rep

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The salivary glands of animals have great potential to act as powerful bioreactors to produce human therapeutic proteins. Human nerve growth factor (hNGF) is an important pharmaceutical protein that is clinically effective in the treatment of many human neuronal and non-neuronal diseases. In this study, we generated 18 transgenic (TG) founder mice each carrying a salivary gland specific promoter-driven hNGF transgene. A TG mouse line secreting high levels of hNGF protein in its saliva (1.36 μg/mL) was selected. hNGF protein was successfully purified from the saliva of these TG mice and its identity was verified. The purified hNGF was highly functional as it displayed the ability to induce neuronal differentiation of PC12 cells. Furthermore, it strongly promoted proliferation of TF1 cells, above the levels observed with mouse NGF. Additionally, saliva collected from TG mice and containing unpurified hNGF was able to significantly enhance the growth of TF1 cells. This study not only provides a new and efficient approach for the synthesis of therapeutic hNGF but also supports the concept that salivary gland from TG animals is an efficient system for production of valuable foreign proteins.

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confidence: 73%

Production of functional human nerve growth factor from the saliva of transgenic mice by using salivary glands as bioreactors

Zeng

Zhu

et al. 2017

Sci Rep

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“…Of them, the highest number of the identified proteins was also quantified in S‐Trap and FASP methods, followed by Acetone‐M method. Out of the 771 proteins, 85 proteins including mammaglobin‐A, olfactomedin‐like protein 3, retinol dehydrogenases 10 and 14, were first identified in cow MFGM fraction (as shown Supporting Information Table 4), compared to the previously identified proteins of the pasteurized milk, in which 74 cow MFGM proteins were identified by shotgun proteomic method , and of raw milk in recent studies . In raw milk, 554 proteins were identified and quantified by LFQ approach, and 520 proteins quantified by iTRAQ proteomics approach in bovine MFGM fraction .…”

Section: Resultsmentioning

confidence: 97%

Evaluation of six sample preparation procedures for qualitative and quantitative proteomics analysis of milk fat globule membrane

2018

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Proteomic analysis of membrane proteins is challenged by the proteins solubility and detergent incompatibility with MS analysis. No single perfect protocol can be used to comprehensively characterize the proteome of membrane fraction. Here, we used cow milk fat globule membrane (MFGM) proteome analysis to assess six sample preparation procedures including one in-gel and five in-solution digestion approaches prior to LC-MS/MS analysis. The largest number of MFGM proteins were identified by suspension trapping (S-Trap) and filter-aided sample preparation (FASP) methods, followed by acetone precipitation without clean-up of tryptic peptides method. Protein identifications with highest average coverage was achieved by Chloroform/MeOH, in-gel and S-Trap methods. Most distinct proteins were identified by FASP method, followed by S-Trap. Analyses by Venn diagram, principal-component analysis, hierarchical clustering and the abundance ranking of quantitative proteins highlight differences in the MFGM fraction by the all sample preparation procedures. These results reveal the biased proteins/peptides loss occurred in each protocol. In this study, we found several novel proteins that were not observed previously by in-depth proteomics characterization of MFGM fraction in milk. Thus, a combination of multiple procedures with orthologous properties of sample preparation was demonstrated to improve the protein sequence coverage and expression level accuracy of membrane samples.

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“…In particular, reversed-phase high-performance liquid chromatography (RP-HPLC) is a common analytical method that has been widely used. Previously, liquid chromatography and mass spectrometry were applied to measure human OPN only as a biomarker [9], for identification and characterization of bovine OPN fragments [10,11] and to separate N-and C-terminal in investigating the binding of human OPN to integrin [12,13] as summarized in Table 1 (Section 3.1).…”

Section: Introductionmentioning

confidence: 99%

A Reversed-Phase HPLC Method for Determination of Osteopontin in Infant Formula

Wazed

Farid

2019

Applied Sciences

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Osteopontin (OPN) is a multifunctional whey protein which has recently received much attention for possibly applications in fortifying infant milk formula (IMF) with its bioactivity. However, to date, there is no established high-performance liquid chromatography (HPLC) method to quantify this protein in milk or IMF. In this study, a rapid, simple, isocratic and reliable reversed-phase HPLC method was developed and validated to quantify the OPN in IMF. A C18 column (4.6 × 150 mm × 5 micron) was employed with 20% of 0.1% trifluoroacetic acid (TFA) and 80% of 60% acetonitrile in 0.1% TFA for 10 min detected at 214 nm. The flow rate was 0.3 mL/min with an injection volume of 10 µL. The column temperature was 40 °C, and the peak appeared after 4 min. The validation was based on the system suitability, linearity (r2 = 0.999), limit of detection (LOD) (0.14 mg/L), limit of quantitation (LOQ) (0.41 mg/L), precision (% relative standard deviation (RSD) < 0.2), recovery (% RSD < 3) and robustness. The results confirm that the method developed is suitable for OPN determination in IMF.

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Characterization of the bovine milk proteome in early-lactation Holstein and Jersey breeds of dairy cows

Cited by 68 publications

References 57 publications

Production of functional human nerve growth factor from the saliva of transgenic mice by using salivary glands as bioreactors

Production of functional human nerve growth factor from the saliva of transgenic mice by using salivary glands as bioreactors

Evaluation of six sample preparation procedures for qualitative and quantitative proteomics analysis of milk fat globule membrane

A Reversed-Phase HPLC Method for Determination of Osteopontin in Infant Formula

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