scholarly journals Marked difference in efficiency of the digestive enzymes pepsin, trypsin, chymotrypsin, and pancreatic elastase to cleave tightly folded proteins

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Zhirong Fu ◽  
Srinivas Akula ◽  
Michael Thorpe ◽  
Lars Hellman
Keyword(s):  
Amino Acids ◽  
Intestinal Mucosa ◽  
Serine Proteases ◽  
Major Effect ◽  
Low Ph ◽  
Pancreatic Enzymes ◽  
Short Peptides ◽  
Pancreatic Elastase ◽  
Native Proteins ◽  
Folded Proteins

Abstract In order for the intestinal mucosa to absorb dietary proteins they have to be digested into single amino acids or very short peptides of a length of not more than four amino acids. In order to study the efficiency of the digestive endopeptidases to digest folded proteins we have analyzed several target proteins under different conditions, native proteins, heat denatured and acid treated. The three pancreatic serine proteases, trypsin, chymotrypsin, and pancreatic elastase, were found to be remarkable inefficient in cleaving native folded proteins whereas pepsin, which acts at a very low pH (pH 1.2) was much more efficient, possibly due to the denaturing conditions and thereby better accessibility to internal cleavage sites at the low pH. Heat treatment improved the cleavage considerably by all three pancreatic enzymes, but acid treatment followed by return to neutral pH did not have any major effect. Cleavage at the low pH when the protein is in a denatured state, is apparently very efficient. This indicates that pepsin is the prime enzyme cleaving the properly folded native proteins and that the pancreatic enzymes primarily are involved in generating single amino acids or very short peptides for efficient uptake by the intestinal mucosa.

scholarly journals Identification of potential autoantigens in anti-CCP-positive and anti-CCP-negative rheumatoid arthritis using citrulline-specific protein arrays

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Thomas B. G. Poulsen ◽  
Dres Damgaard ◽  
Malene M. Jørgensen ◽  
Ladislav Senolt ◽  
Jonathan M. Blackburn ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Protein Microarray ◽  
Specific Protein ◽  
Immune Reactivity ◽  
Igg Antibodies ◽  
Native Proteins ◽  
Array Technology ◽  
Citrullinated Proteins ◽  
Folded Proteins

AbstractThe presence or absence of autoantibodies against citrullinated proteins (ACPAs) distinguishes two main groups of rheumatoid arthritis (RA) patients with different etiologies, prognoses, disease severities, and, presumably, disease pathogenesis. The heterogeneous responses of RA patients to various biologics, even among ACPA-positive patients, emphasize the need for further stratification of the patients. We used high-density protein array technology for fingerprinting of ACPA reactivity. Identification of the proteome recognized by ACPAs may be a step to stratify RA patients according to immune reactivity. Pooled plasma samples from 10 anti-CCP-negative and 15 anti-CCP-positive RA patients were assessed for ACPA content using a modified protein microarray containing 1631 different natively folded proteins citrullinated in situ by protein arginine deiminases (PADs) 2 and PAD4. IgG antibodies from anti-CCP-positive RA plasma showed high-intensity binding to 87 proteins citrullinated by PAD2 and 99 proteins citrullinated by PAD4 without binding significantly to the corresponding native proteins. Curiously, the binding of IgG antibodies in anti-CCP-negative plasma was also enhanced by PAD2- and PAD4-mediated citrullination of 29 and 26 proteins, respectively. For only four proteins, significantly more ACPA binding occurred after citrullination with PAD2 compared to citrullination with PAD4, while the opposite was true for one protein. We demonstrate that PAD2 and PAD4 are equally efficient in generating citrullinated autoantigens recognized by ACPAs. Patterns of proteins recognized by ACPAs may serve as a future diagnostic tool for further subtyping of RA patients.

scholarly journals Comparison of liver glycosylasparaginases from six vertebrates

1992 ◽  
Vol 282 (3) ◽  
pp. 891-897 ◽  
Author(s):  
O K Tollersrud ◽  
N N Aronson
Keyword(s):  
Amino Acids ◽  
Reaction Mechanism ◽  
Basic Structure ◽  
Low Ph ◽  
Beta Subunit ◽  
Beta Subunits ◽  
Species Identity ◽  
Closely Related Species ◽  
Human Enzyme ◽  
Specific Antisera

Structural and physical properties of glycosylasparaginase (EC 3.5.1.26) from the livers of human, pig, cow, rat, mouse and chicken were compared. The enzyme in all species had a common basic structure of two N-glycosylated subunits of about 24 (alpha) and 20 (beta) kDa joined by non-covalent forces. Subunit-specific antisera against the rat glycosylasparaginase bound specifically and sensitively to the corresponding subunits from all species. Identity of 80% of the amino acids was found between the N-terminal sequences of corresponding pig and rat glycosylasparaginase alpha- and beta-subunits and the deduced sequence from a human glycosylasparaginase cDNA [Fisher, Tollersrud & Aronson (1990) FEBS Lett. 269, 440-444]. The beta-subunit from all three species has an N-terminal threonine reported to be involved in the reaction mechanism for the human enzyme [Kaartinen, Williams, Tomich, Yates, Hood & Mononen (1991) J. Biol. Chem. 266, 5860-5869]. The native enzyme appeared as a heterodimer among the mammals, whereas the chicken enzyme had a greater molecular mass and is probably either a tetramer or a heterodimer bound to an unrelated peptide(s). All glycosylasparaginases were thermostable, requiring temperatures between 65 degrees C and 80 degrees C to be irreversibly inactivated. In addition, they were unusually stable at high pH and remained active in the presence of SDS except at low pH. The pH maximum was between 5.5 and 6 except for the rat and mouse enzymes which had a broad maximum between pH 7 and 8. A number of other properties were observed which also distinguish the enzyme from individual and closely related species.

The Native Proteins as Polycondensations of Amino Acids

Science ◽  
1948 ◽  
Vol 107 (2783) ◽  
pp. 445-446
Author(s):  
Dorothy Wrinch
Keyword(s):  
Amino Acids ◽  
Native Proteins

scholarly journals Accumulation of amino acids by the perfused rat liver in the presence of ethanol

1973 ◽  
Vol 134 (3) ◽  
pp. 697-705 ◽  
Author(s):  
Hans A. Krebs ◽  
Reginald Hems ◽  
Patricia Lund
Keyword(s):  
Amino Acids ◽  
Rat Liver ◽  
Major Effect ◽  
Major Product ◽  
Control Mechanisms ◽  
Perfused Rat Liver ◽  
Carbamoyl Phosphate ◽  
Lactate Formation ◽  
Gluconeogenesis From Alanine ◽  
Almost All

1. The rate of gluconeogenesis from alanine in the perfused rat liver is affected by the presence of other metabolizable substances, especially fatty acids, ornithine and ethanol. Gluconeogenesis is accelerated by oleate and by ornithine. When both oleate and ornithine were present the acceleration was greater than expected on the basis of mere additive effects. 2. Much NH3 and some urea were formed from alanine when no ornithine was added. With ornithine almost all the nitrogen released from alanine appeared as urea. 3. Lactate was a major product of alanine metabolism. Addition of oleate, and especially of oleate plus ornithine, decreased lactate formation. 4. Ethanol had no major effect on gluconeogenesis from alanine when this was the sole added precursor. Gluconeogenesis was strongly inhibited (87%) when oleate was also added, but ethanol greatly accelerated gluconeogenesis when ornithine was added together with alanine. 5. In the absence of ethanol the alanine carbon and alanine nitrogen removed were essentially recovered in the form of glucose, lactate, pyruvate, NH3 and urea. 6. In the presence of ethanol the balance of both alanine carbon and alanine nitrogen showed substantial deficits. These deficits were largely accounted for by the formation of aspartate and glutamine, the formation of which was increased two- to three-fold. 7. When alanine was replaced by lactate plus NH4Cl, ethanol also caused a major accumulation of amino acids, especially of aspartate and alanine. 8. Earlier apparently discrepant results on the effects of ethanol on gluconeogenesis from alanine are explained by the fact that under well defined conditions ethanol can inhibit, or accelerate, or be without major effect on the rate of gluconeogenesis. 9. It is pointed out that in the synthesis of urea through the ornithine cycle half of the nitrogen must be supplied in the form of asparate and half in the form of carbamoyl phosphate. The accumulation of aspartate and other amino acids suggests that ethanol interferes with the control mechanisms which regulate the stoicheiometric formation of aspartate and carbamoyl phosphate.

scholarly journals Analysis of Htra Gene from Zebrafish (Danio Rerio)

2015 ◽  
Vol 10 (2) ◽  
Author(s):  
M. Murwantoko ◽  
Chio Oka ◽  
Masashi Kawaichi
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Danio Rerio ◽  
Serine Proteases ◽  
Catalytic Domain ◽  
Fruit Fly ◽  
Wide Range ◽  
Igf Binding ◽  
Bacterial Homolog

HtrA which is characterized by the combination of a trypsin-like catalytic domain with at least one C-terminalPDZ domain is a highly conserved family of serine proteases found in a wide range of organisms. However theidentified HtrA family numbers varies among spesies, for example the number of mammalian, Eschericia coli,fruit fly-HtrA family are 4, 3 and 1 gene respectively. One gene is predicted exist in zebrafish. Since no completeinformation available on zebrafish HtrA, in this paper zebrafish HtrA (zHtrA) gene was analyzed. The zHtrA isbelonged to HtrA1 member and predicted encodes 478 amino acids with a signal peptide, a IGF binding domain,a Kazal-type inhibitor domain in the up stream of HtrA-bacterial homolog. At the amino acid sequence the zHtrA1showed the 69%, 69%, 68%, 54% and 54% with the rat HtrA1, mouse HtrA1, human HtrA1, human HtrA3 andmouse HtrA4 respectively. The zHtrA1 is firstly expressed at 60 hpf and mainly in the vertebral rudiments in thetail region.

Biochemical and functional characterization of Mycobacterium tuberculosis ketol-acid reductoisomerase

Microbiology ◽  
2021 ◽  
Vol 167 (9) ◽  
Author(s):  
Nirbhay Singh ◽  
Anu Chauhan ◽  
Ram Kumar ◽  
Sudheer Kumar Singh
Keyword(s):  
Amino Acids ◽  
Mycobacterium Tuberculosis ◽  
Type Species ◽  
Low Ph ◽  
Stress Conditions ◽  
Starvation Stress ◽  
Content Type ◽  
E Coli ◽  
Link Type

Branched-chain amino acids (BCAAs) are essential amino acids, but their biosynthetic pathway is absent in mammals. Ketol-acid reductoisomerase (IlvC) is a BCAA biosynthetic enzyme that is coded by Rv3001c in Mycobacterium tuberculosis H37Rv (Mtb-Rv) and MRA_3031 in M. tuberculosis H37Ra (Mtb-Ra). IlvCs are essential in Mtb-Rv as well as in Escherichia coli . Compared to wild-type and IlvC-complemented Mtb-Ra strains, IlvC knockdown strain showed reduced survival at low pH and under low pH+starvation stress conditions. Further, increased expression of IlvC was observed under low pH and starvation stress conditions. Confirmation of a role for IlvC in pH and starvation stress was achieved by developing E. coli BL21(DE3) IlvC knockout, which was defective for growth in M9 minimal medium, but growth could be rescued by isoleucine and valine supplementation. Growth was also restored by complementing with over-expressing constructs of Mtb-Ra and E. coli IlvCs. The E. coli knockout also had a survival deficit at pH=5.5 and 4.5 and was more susceptible to killing at pH=3.0. The biochemical characterization of Mtb-Ra and E. coli IlvCs confirmed that both have NADPH-dependent activity. In conclusion, this study demonstrates the functional complementation of E. coli IlvC by Mtb-Ra IlvC and also suggests that IlvC has a role in tolerance to low pH and starvation stress.

The Enhancement of Iron Absorption in Rats by D-Sorbitol

PEDIATRICS ◽  
1958 ◽  
Vol 22 (4) ◽  
pp. 705-705
Keyword(s):  
Amino Acids ◽  
Vitamin B12 ◽  
Intestinal Mucosa ◽  
Iron Absorption ◽  
Significant Enhancement ◽  
Clinical Therapy ◽  
Tartaric Acids

Recently investigations have shown that the carbohydrate, D-Sorbitol, enhances the absorption of vitamin B12. Certain amino acids and ascorbic, citric and tartaric acids tend to enhance iron absorption. In view of these findings, the authors undertook to determine the effect of D-sorbitol on iron absorption in the rat. It was found that D-sorbitol causes significant enhancement of the absorption of iron in iron-depleted as well as in normal rats. It was also shown that the amount of iron absorbed increased in proportion to the amount of D-sorbitol administered with the iron. These experiments suggest that the absorption of iron may not be controlled exclusively by the amount of apoferritin in the intestinal mucosa. Possible beneficial or deleterious consequences of the application of these findings to clinical therapy remain to be determined.

scholarly journals The Glycine- and Proline-Rich Protein AtGPRP3 Negatively Regulates Plant Growth in Arabidopsis

2020 ◽  
Vol 21 (17) ◽  
pp. 6168
Author(s):  
Xiaojing Liu ◽  
Xin Wang ◽  
Xin Yan ◽  
Shaobo Li ◽  
Hui Peng
Keyword(s):  
Amino Acids ◽  
Plant Growth ◽  
Physiological Role ◽  
Short Peptides ◽  
Hydrophobic Domain ◽  
Gfp Fusion Protein ◽  
Repeat Domain ◽  
Arabidopsis Plant ◽  
Proline Rich Protein

Glycine- and proline-rich proteins (GPRPs) comprise a small conserved family that is widely distributed in the plant kingdom. GPRPs are relatively short peptides (<200 amino acids) that contain three typical domains, including an N-terminal XYPP-repeat domain, a middle hydrophobic domain rich in alanine, and a C-terminal HGK-repeat domain. These proteins have been proposed to play fundamental roles in plant growth and environmental adaptation, but their functions remain unknown. In this study, we selected an Arabidopsis GPRP (AtGPRP3) to profile the physiological role of GPRPs. Transcripts of AtGPRP3 could be detected in the whole Arabidopsis plant, but greater amounts were found in the rosette, followed by the cauline. The AtGPRP3::GFP fusion protein was mainly localized in the nucleus. The overexpression and knockout of AtGPRP3, respectively, retarded and accelerated the growth of Arabidopsis seedlings, while the increase in the growth rate of atgprp3 plants was offset by the complementary expression of AtGPRP3. CAT2 and CAT3, but not CAT1, interacted with AtGPRP3 in the nuclei of Arabidopsis protoplasts. The knockout of CAT2 by CRISPR-Cas9 retarded the growth of the Arabidopsis seedlings. Together, our data suggest that AtGPRP3 negatively regulates plant growth, potentially through CAT2 and CAT3.

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