• Journal of Microbiology and Biotechnology
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• v.27 no.2
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• pp.277-288
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• 2017

Rhizomucor miehei NRRL 5282 and Rhizopus oryzae NRRL 1526 can produce lipases with high synthetic activities in wheat bran-based solid-state culture. In this study, the purification and biochemical characterization of the lipolytic activities of these lipases are presented. SDS-PAGE indicated a molecular mass of about 55 and 35 kDa for the purified R. miehei and Rh. oryzae enzymes, respectively. p-Nitrophenyl palmitate (pNPP) hydrolysis was maximal at $40^{\circ}C$ and pH 7.0 for the R. miehei lipase, and at $30^{\circ}C$ and pH 5.2 for the Rh. oryzae enzyme. The enzymes showed almost equal affinity to pNPP, but the $V_{max}$ of the Rh. oryzae lipase was about 1.13 times higher than that determined for R. miehei using the same substrate. For both enzymes, a dramatic loss of activity was observed in the presence of 5 mM $Hg^{2+}$, $Zn^{2+}$, or $Mn^{2+}$, 10 mM N-bromosuccinimide or sodium dodecyl sulfate, and 5-10% (v/v) of hexanol or butanol. At the same time, they proved to be extraordinarily stable in the presence of n-hexane, cyclohexane, n-heptane, and isooctane. Moreover, isopentanol up to 10% (v/v) and propionic acid in 1 mM concentrations increased the pNPP hydrolyzing activity of R. miehei lipase. Both enzymes had 1,3-regioselectivity, and efficiently hydrolyzed p-nitrophenyl (pNP) esters with C8-C16 acids, exhibiting maximum activity towards pNP-caprylate (R. miehei) and pNP-dodecanoate (Rh. oryzae). The purified lipases are promising candidates for various biotechnological applications.

• Journal of the Korean Applied Science and Technology
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• v.18 no.4
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• pp.306-315
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• 2001

Our study is aimed at proposal of systematic verification method of molecular structure using measuring method of selective ionic determination and spectrometry on 34 kinds of surfactants such as sodium dodecyl sulfate(SDS) which are most widely used today. In the IR spectrum, unsaturated fatty acids reveal themselves by HC= at $3000{\sim}3020cm^{-1}$, and intensity of $720cm^{-1}$ depends on carbon length of alkyl group. Also ethylene oxide(EO) adducts exhibit weak characteristic bands by $-CH_{2}-CH_{2}-O$ at 1350, 1100 and $950cm^{-1}$. Isethionate can be distinguished from diester succinate by intensity ratio of 1740 and $1200cm^{-1}$ spectrums, the ratio of latter is close to 1 due to 2 carboxylate radical in diester succinate. Quaternary ammonium salts exhibit characteristic band of $C_{4}N^{+}$ at $1000-900㎝^{-1}$. In the case of dialkyl dimethyl ammonium salts in quaternary ammonium surfactants, the spectrum of $3000cm^{-1}$ by $N-CH_{3}$ collapses to a very weak band at $3020cm^{-1}$. In ammonium heterocyclic derivatives, pyridinium salts show characteristic bands at 1640 and $1460cm^{-1}$, while imidazolinium salts exhibit characteristic band at $1620-1610cm^{-1}$. In the characteristic spectrum at $1080-1050cm^{-1}$ on OH radicals of the alkyl esters, primary alcohol appears as weak band and the 2 bands show in almost same intensity when primary and secondary alcohols exist together in one molecule. Also, alkyl ester of polyhydric alcohols appears as various broad band.

• Journal of the Korean Society of Food Science and Nutrition
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• v.18 no.3
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• pp.287-299
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• 1989

This research was conducted in order to investigate thermal stability and nutritional value of corn lipids and proteins during thermal processing. The lipids of raw and popped corn were fractionated and analyzed by column and gas chromatography. The effect of thermal processing on corn proteins was examined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and amino acid analysis. There was no remarkable change in proximate compositions during thermal processing. The lipid fractions obtained by silicic acid column chromatography were composed of neutral lipid(93.5%), glycolipid(3.8%), and phospholipid(2.7%), Although the thermal processing showed the increase in the ratio of unsaturated/saturated fatty acid, there was no significant differences in the lipid composition between raw and popped corn. Most of each protein fractions had lower molecular weight than 25,000 dalton and albumin fractions were distributed in the molecular weight range 11,500-94,000 daltons. Popped corn proteins did not show marked differences in their electrophoretic migrations when compared with raw corn proteins.

• Journal of the Korean Society of Food Science and Nutrition
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• v.38 no.6
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• pp.801-806
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• 2009

A species producing transglutaminase (EC 2.3.2.13) was isolated from forest soil and identified as Streptomyces platensis YK-2. The transglutaminase was purified from culture broth by 50% methanol precipitation, followed by successive chromatography on DEAE-Sephadex. The yield and purification-fold was 63.4% and 2.2-fold, respectively. The purified microbial transglutaminase (MTG) migrated as a single band of approximately 45 kDa upon sodium dodecyl sulfate polyacrylamide gel eletrophoresis. The isoelectric point determined by multichambered electrofocusing was pH $6.0{\sim}7.0$. The enzyme was strongly inhibited by $Hg^{++}$, but was activated by $Cd^{++}$, $Mg^{++}$, $Mn^{++}$, $Pb^{++}$ and reducing agents such as dithiothreitol and mercaptoethanol.

• Korean Journal of Agricultural Science
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• v.45 no.4
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• pp.635-643
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• 2018

Bovine milk is widely consumed by humans and is a primary ingredient of dairy foods. Proteomic approaches have the potential to elucidate complex milk proteins and have been used to study milk of various species. Here, we performed a proteomic analysis using 2-dimensional electrophoresis (2-DE) and matrix assisted laser desorption ionization-time of flight mass spectrometer (MALDI-TOF MS) to identify whey proteins in bovine milk obtained soon after parturition (bovine early milk). The major casein proteins were removed, and the whey proteins were analyzed with 2-dimensional polyacrylamide gel electrophoresis (2-D PAGE). The whey proteins (2 mg) were separated by pI and molecular weight across pH ranges of 3.0 - 10.0 and 4.0 - 7.0. The 2-DE gels held about 300 to 700 detectable protein spots. We randomly picked 12 and nine spots that were consistently expressed in the pH 3.0 - 10.0 and pH 4.0 - 7.0 ranges, respectively. Following MALDI-TOF MS analysis, the 21 randomly selected proteins included proteins known to be present in bovine milk, such as albumin, lactoferrin, serum albumin precursor, T cell receptor, polymeric immunoglobulin receptor, pancreatic trypsin inhibitor, aldehyde oxidase and microglobulin. These proteins have major functions in immune responses, metabolism and protein binding. In summary, we herein identified both known and novel whey proteins present in bovine early milk, and our sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis revealed their expression pattern.

• Fisheries and Aquatic Sciences
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• v.24 no.12
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• pp.406-414
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• 2021

The objective of this study was to investigate the yield and characteristics of collagen protein extracted from the muscle of four different species of mantis shrimp: Miyakella nepa, Harpiosquilla harpax, Erugosquilla woodmasoni, and Odontodactylus cultrifer. Mantis shrimp muscle was extracted by using a pepsin-solubilization technique, with 0.5 M acetic acid and 5% pepsin enzyme. The highest collagen yield was from M. nepa muscle (0.478 ± 0.06%), which was significantly greater (p < 0.05) than that from H. harpax, O. cultrifer, and E. woodmasoni (0.313 ± 0.03%, 0.123 ± 0.02%, and 0.015 ± 0.00%, respectively). The freeze-dried collagen appeared as thin fibers, and formed an opaque film. The pepsin-soluble collagen (PSC) from four mantis shrimp species was analyzed by gel electrophoresis. The results showed that all species of mantis shrimp contained type I collagen, consisting of β, α1, and α2 subunits with average molecular weights of 250, 145, and 118 kDa, respectively. The study of the solubility of collagen showed that, for NaCl, collagen had the highest relative solubility in 2% NaCl (80.20 ± 4.95%). In contrast, the solubility decreased at higher NaCl concentrations. However, in terms of pH, collagen had the highest relative solubility at pH 3 (91.32 ± 5.14%), and its solubility decreased at higher pH. FT-IR spectroscopy was used to compare the collagen with a model compound. Five wavenumbers in the spectrum for model collagen were identified: Amide A (3,406-3,421 cm^-1), amide B (2,916-2,940 cm^-1), amide I (1,639-1,640 cm^-1), amide II (1,539-1,570 cm^-1), and amide III (1,234-1,250 cm^-1).

• International Journal of Industrial Entomology and Biomaterials
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• v.45 no.2
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• pp.93-98
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• 2022

To assess the feasibility of silk sericin for non-textile application, the storage stability and biological safety of sericin were examined. It was extracted at 37℃, 70℃, 100℃, and 121℃ for 1, 3, and 5 h to elucidate the effect of extraction condition on the stability and safety of silk sericin. The solubility was increased till approximately 26% with extraction temperature of 121℃ for 1 h. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that the molecular weight distribution depended on the extraction conditions. Extracted sericin displayed typical UV absorption bands upon spectrometric analysis. To examine the reproducibility of its obtained conformation, sericin was extracted thrice and its circular dichroism (CD) spectra was measured each time. Most CD spectra showed reproducibility regardless of temperature and time except under 100℃ extraction condition. The diversity of CD spectrum showed gradual reduction and was finally coincident with extraction time from 1 to 5 h. Notably, sericin has a negative peak of approximately 200 nm attributed to random coil conformation, regardless of extraction condition. However, at the 100℃ extraction condition, sericin showed both bands to be negative bands of approximately 200 and 220 nm, respectively. Sericin was centrifuged to determine the stability of storage conditions. The sericin extracted at 100℃ and 121℃ for 1 h was found to form gel rapidly within 1 h, but at 121℃ condition, the gel fraction was approximately 20% within 1 h which retained its phase regardless of storage time. The gel fraction of sericin extracted at 100℃ for 5 h increased with time, however at the 121℃ for 5 h condition, the gel fraction was measured to be less than 10% regardless of increase in storage time. Petriflim^TM AC plates test showed that sericin was safe from aerobic bacteria activity by extraction under high temperature.

• Food Science of Animal Resources
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• v.28 no.2
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• pp.154-159
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• 2008

To investigate the rheological properties of protein mixed gels mediated by microbial transglutaminase (MTGase), pork myofibrillar protein (MFP), sodium caseinate (SC) and their mixture (MS), the various gels were incubated at different temperatures for various times. Extracted MFP, SC and their mixture (MS, 1:1) were incubated at different temperatures ($4^{\circ}C$ vs $37^{\circ}C$) for various times (0, 0.5, 2, 4 hr), and assessed for viscosity, gel strength and other characteristics using differential scanning calorimeter (DSC) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). DSC measurements showed that incubation at $37^{\circ}C$ rather than $4^{\circ}C$ caused marked changes in thermal transition, and MS displayed similar thermal curves (three endothermic transitions) to MFP and SC alone. After incubation at $37^{\circ}C$ for 2 hrs, the viscosity (cP) of MS increased (p<0.05) due to induction by MTGase, whereas no differences were observed at $4^{\circ}C$. However, gel strength values were no different, regardless of incubation temperatures and times. Future research will address how longer incubation times affect the functionality of protein mixed gels mediated by MTGase.

• Journal of the Korean Society of Food Science and Nutrition
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• v.33 no.10
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• pp.1668-1675
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• 2004

The effect of pH on surface hydrophobicity, sulfhydryl group, infrared spectrum, SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) pattern and enthalpy was investigated in recovered protein from mackerel and frozen blackspotted croaker by alkaline processing. Hydrophobic residue in myofibrillar protein exposed to the surface of protein, and hydrophobic interaction were the highest around 6$0^{\circ}C$. The surface hydrophobicity was different between myofibrillar protein and myofibrillar protein including sarcoplasmic protein (recovered protein). The peak at 1636 c $m^{-l}$ was increased with pH, and the recovered protein was unfolded in alkali pH. Difference of surface and total sulfhydryl group at pH 7.0 and 10 was comparative high, and decrease of surface sulfhydryl group indicated formation of S-S bonds. Mackerel and frozen blackspotted croaker in alkaline pH showed bands of polymerized myosin heavy chain on SDS-PAGE pattern. The transition temperatures of recovered protein were 33.1, 44.3 and 65.5$^{\circ}C$. Gelation of recovered protein from alkali processing was estimated by increase of $\beta$-sheet structure by pH treatment, S-S bonds by oxidation of surface sulfhydryl group in heating, polymerization of myosin heavy chain in order.r.

• Asian-Australasian Journal of Animal Sciences
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• v.22 no.4
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• pp.534-541
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• 2009

The aim of this study was to evaluate the effects of gamma irradiation (${\gamma}$-irradiation) at doses of 15, 30 and 45 kGy on chemical composition, anti-nutritional factors, ruminal dry matter (DM) and crude protein (CP) degradibility, in vitro CP digestibility and to monitor the fate of true proteins of full-fat soybean (SB) in the rumen. Nylon bags of untreated or ${\gamma}$-irradiated SB were suspended in the rumens of three ruminally-fistulated bulls for up to 48 h and resulting data were fitted to a nonlinear degradation model to calculate degradation parameters of DM and CP. Proteins of untreated and treated SB bag residues were fractionated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Digestibility of rumen undegraded CP was estimated using the three-step in vitro procedure. The chemical composition of raw and irradiated soybeans was similar. Results showed that phytic acid in ${\gamma}$-irradiated SB at dose of 30 kGy was eliminated completely. The trypsin inhibitor activity of 15, 30 and 45 kGy ${\gamma}$-irradiated SB was decreased (p<0.01) by 18.4, 55.5 and 63.5%, respectively. From in sacco results, ${\gamma}$-irradiation decreased (p<0.05) the washout fractions of DM and CP at doses of 30 and 45 kGy, but increased (p<0.05) the potentially degradable fractions. Gamma irradiation at doses of 15, 30 and 45 kGy decreased (p<0.05) effective degradability of CP at a rumen outflow rate of 0.05 $h^{-1}$ by 4.4, 14.4 and 26.5%, respectively. On the contrary, digestibility of ruminally undegraded CP of irradiated SB at doses of 30 and 45 kGy was improved (p<0.05) by 12 and 28%, respectively. Electrophoretic analysis of untreated soybean proteins incubated in the rumen revealed that ${\beta}$-conglycinin subunits had disappeared at 2 h of incubation time, whereas the subunits of glycinin were more resistant to degradation until 16 h of incubation. From the SDS-PAGE patterns, acidic subunits of 15, 30 and 45 kGy ${\gamma}$-irradiated SB disappeared after 8, 8 and 16 h of incubation, respectively, while the basic subunits of glycinin were not degraded completely until 24, 48 and 48 h of incubation, respectively. It was concluded that ${\gamma}$-irradiated soybean proteins at doses higher than 15 kGy could be effectively protected from ruminal degradation.