scholarly journals The controlling effect of carbohydrate in human, rabbit and bovine immunoglobulin G on proteolysis by papain

1969 ◽  
Vol 111 (4) ◽  
pp. 473-478 ◽  
Author(s):  
R. B. Payne
Keyword(s):  
Immunoglobulin G ◽  
Polypeptide Chain ◽  
Initial Rate ◽  
Hinge Region ◽  
Fc Fragment ◽  
Rabbit Igg ◽  
Initial Cleavage ◽  
Bovine Immunoglobulin ◽  
Hydrolysis Of ◽  
Papain Digestion

About two-thirds of the hexose of human and rabbit immunoglobulin G (IgG) was located in the Fc fragment and one-third in the ‘hinge’ region of the γ (heavy) polypeptide chain at the junction of the Fab and Fc fragments. In contrast, bovine IgG contained more hexose in the ‘hinge’ region than in the Fc fragment. The initial cleavage of susceptible IgG molecules into Fab and Fc fragments by papain under the conditions given by Porter (1959) had reached completion after digestion for 2hr., though bovine IgG was digested somewhat more slowly than human or rabbit IgG. The release of ‘hinge’ peptides from human and rabbit IgG had also reached completion by 2hr., but was slower from bovine IgG and continued for several hours longer. Since bovine IgG molecules contained on the average a greater amount of hexose in the ‘hinge’ region, carbohydrate on this part of the γ-chain may influence not only the initial rate of enzymic hydrolysis into Fab and Fc fragments, but also, and to a greater extent, the rate of further limited hydrolysis of the N-terminal regions of the Fc fragment. The presence of carbohydrate in the ‘hinge’ region does not appear to account for the resistance of some IgG molecules to papain digestion and of some Fc fragments to N-terminal degradation.

scholarly journals Leporid immunoglobulin G shows evidence of strong selective pressure on the hinge and CH3 domains

Open Biology ◽  
2014 ◽  
Vol 4 (9) ◽  
pp. 140088 ◽  
Author(s):  
Ana Pinheiro ◽  
Jenny M. Woof ◽  
Tereza Almeida ◽  
Joana Abrantes ◽  
Paulo C. Alves ◽  
...  
Keyword(s):  
Positive Selection ◽  
Immunoglobulin G ◽  
Half Life ◽  
Selective Pressure ◽  
Hinge Region ◽  
European Rabbit ◽  
Rabbit Igg ◽  
Immunological Research ◽  
Constant Domains ◽  
Positively Selected Sites

Immunoglobulin G (IgG) is the predominant serum immunoglobulin and has the longest serum half-life of all the antibody classes. The European rabbit IgG has been of significant importance in immunological research, and is therefore well characterized. However, the IgG of other leporids has been disregarded. To evaluate the evolution of this gene in leporids, we sequenced the complete IGHG for six other genera: Bunolagus , Brachylagus , Lepus , Pentalagus , Romerolagus and Sylvilagus . The newly sequenced leporid IGHG gene has an organization and structure similar to that of the European rabbit IgG. A gradient in leporid IgG constant domain diversity was observed, with the CH1 being the most conserved and the CH3 the most variable domain. Positive selection was found to be acting on all constant domains, but with a greater incidence in the CH3 domain, where a cluster of three positively selected sites was identified. In the hinge region, only three polymorphic positions were observed. The same hinge length was observed for all leporids. Unlike the variation observed for the European rabbit, all 11 Lepus species studied share exactly the same hinge motif, suggesting its maintenance as a result of an advantageous structure or conformation.

scholarly journals Intracellular distribution and degradation of immunoglobulin G and immunoglobulin G fragments injected into HeLa cells.

1983 ◽  
Vol 96 (2) ◽  
pp. 338-346 ◽  
Author(s):  
T McGarry ◽  
R Hough ◽  
S Rogers ◽  
M Rechsteiner
Keyword(s):  
Hela Cells ◽  
Immunoglobulin G ◽  
Protein Modification ◽  
Hinge Region ◽  
Myeloma Protein ◽  
Thin Sections ◽  
Intracellular Degradation ◽  
Rate Limiting Step ◽  
Rabbit Igg ◽  
Intact Rabbit

Intact rabbit immunoglobulin G molecules (IgGs) and their papain or pepsin fragments were radio-iodinated and injected into HeLa cells. Whole IgGs, Fab2, and Fc fragments were degraded with half-lives of 60-90 h, whereas half-lives of Fab fragments were 110 h. These results indicate that proteolytic cleavage in the hinge region of the IgG molecule is not the rate-limiting step in its intracellular degradation. The hingeless human myeloma protein, Mcg, was degraded at the same rate as bulk human IgG, providing further evidence that the proteolytically susceptible hinge region is not important for intracellular degradation of IgG molecules. SDS acrylamide gel analysis of injected rabbit IgG molecules revealed that heavy and light chains were degraded at the same rate. Injected rabbit IgGs and rabbit IgG fragments were also examined on isoelectric focusing gels. Fab, Fab2, and Fc fragments were degraded without any correlation with respect to isoelectric point. Positively charged rabbit IgGs disappeared more rapidly than their negative counterparts, contrary to the trend reported for normal intracellular proteins. The isoelectric points of two mouse monoclonal antibodies were essentially unchanged after injection into HeLa cells, suggesting that the altered isoelectric profile observed for intact rabbit IgG resulted from degradation and not protein modification. The intracellular distributions of IgG fragments and intact rabbit IgG molecules were determined by autoradiography of thin sections through injected cells. Intact IgG molecules were excluded from HeLa nuclei whereas both Fab and Fc fragments readily entered them. Thus, for some proteins, entry into the nuclear compartment is determined primarily by size.

scholarly journals The interaction of protein A and Fc fragment of rabbit immunoglobulin G as probed by complement-fixation and nuclear-magnetic-resonance studies

1977 ◽  
Vol 167 (3) ◽  
pp. 661-668 ◽  
Author(s):  
C Wright ◽  
K J Willan ◽  
J Sjödahl ◽  
D R Burton ◽  
R A Dwek
Keyword(s):  
Nuclear Magnetic Resonance ◽  
Magnetic Resonance ◽  
Immunoglobulin G ◽  
Complement Fixation ◽  
Protein A ◽  
Structural Basis ◽  
Proton Relaxation ◽  
Fc Fragment ◽  
Rabbit Igg ◽  
Nuclear Magnetic

Protein-A-Fc-fragment complexes were observed in sedimentation-velocity experiments by ultracentrifugation. The interaction was studied by protein-fluorescence-quenching titrations of the Fc fragment with protein A, allowing the dissociation constant to be determined under a variety of conditions. The first component of the complement pathway, C1, is activated by complexes of protein A with rabbit IgG (immunoglobulin G), and the structural basis for this interaction was studied by using n.m.r. (nuclear magnetic resonance). The four Fc-fragment binding sites on protein A were shown to contain aromatic amino acids, and to be connected by mobile hydrophilic regions. Neither n.m.r. nor proton-relaxation-enhancement studies show evidence of a large conformational change of the Fc fragment on binding protein A, and this suggests that the cross-linking of the Fc fragments may be primarily responsible for the activation of component C1. This is supported by the inability of a univalent tryptic fragment of protein A to activate complement fixation by rabbit IgG.

scholarly journals Stepwise cleavage of rabbit immunoglobin G by papain and isolation of four types of biologically active Fc fragments

1969 ◽  
Vol 112 (3) ◽  
pp. 343-355 ◽  
Author(s):  
Sayaka Utsumi
Keyword(s):  
Guinea Pig ◽  
Immunoglobulin G ◽  
Hinge Region ◽  
Biologically Active ◽  
Antigenic Structure ◽  
Antigenic Determinants ◽  
Fc Fragment ◽  
Pig Skin ◽  
Neutral Ph ◽  
Immunoglobin G

Four types of Fc fragments of different sizes were isolated by papain treatment of rabbit immunoglobulin G under various conditions and by subsequent chromatographic procedures. 1. Brief digestion at neutral pH without reduction produced a molecule in which the Fab and Fc fragments were still linked by a pair of labile disulphide bridges, and the Fc fragment released by cleaving these bonds, called 1Fc fragment, contained a portion of the ‘hinge’ region including an interchain disulphide bridge. Both complement-binding and guinea-pig skin-binding activities were retained by this fragment, which had mol. wt. 48000. 2. Prolonged digestion at neutral pH of immunoglobulin G whose labile inter-heavy-chain disulphide bridges had been reduced removed the ‘hinge’ region, giving mFc fragments (mol. wt. 46000), which lacked the capacity to bind guinea-pig skin but retained the antigenic as well as the complement-binding activities of 1Fc fragment completely. 3. Digestion at pH5·0 yielded a smaller fragment, sFc (mol. wt. 40000), which was no longer able to bind complement. Though the antigenic structure was intact, sFc fragment was curiously unable to precipitate with antibodies to the N-terminal determinants. 4. Fragment stFc (mol. wt. 25000), representing the C-terminal portion of Fc fragment, was formed from all the larger fragments by digestion at pH4·5. Only the C-terminal antigenic determinants were retained by stFc fragment.

scholarly journals Antigenic Crossreactivity and Lipopolysaccharide Neutralization Properties of Bovine Immunoglobulin G

1995 ◽  
Vol 78 (12) ◽  
pp. 2745-2752 ◽  
Author(s):  
G.M. Tomita ◽  
D.A. Todhunter ◽  
J.S. Hogan ◽  
K.L. Smith
Keyword(s):  
Immunoglobulin G ◽  
Bovine Immunoglobulin

scholarly journals The subunit and polypeptide-chain structure of rabbit secretory immunoglobulin A. Isolation of a proteolytic fragment suitable for sequence studies on the variable region of α-chain

1977 ◽  
Vol 167 (1) ◽  
pp. 245-253 ◽  
Author(s):  
A P Johnstone ◽  
L E Mole
Keyword(s):  
Immunoglobulin G ◽  
Polypeptide Chain ◽  
Immunoglobulin A ◽  
Variable Region ◽  
Chain Structure ◽  
Secretory Immunoglobulin A ◽  
Primary Sequence ◽  
Relative Resistance ◽  
Proteolytic Fragment ◽  
Secretory Immunoglobulin

A method was developed for the preparation of a proteolytic fragment of rabbit secretory immunoglobulin A (sIgA) which contains the variable region of the alpha-chain; this fragment is suitable for primary-sequence studies. The serologically defined subclasses of sIgA are shown to correlate partially with the nature of the binding of a constituent chain of sIgA, called secretory piece. Data are also presented on the relative resistance of sIgA to enzymic and reductive cleavage, compared with immunoglobulin G.

scholarly journals Backbone 1H, 13C, and 15N resonance assignments of the Fc fragment of human immunoglobulin G glycoprotein

2014 ◽  
Vol 9 (2) ◽  
pp. 257-260 ◽  
Author(s):  
Hirokazu Yagi ◽  
Ying Zhang ◽  
Maho Yagi-Utsumi ◽  
Takumi Yamaguchi ◽  
Shigeru Iida ◽  
...  
Keyword(s):  
Immunoglobulin G ◽  
Resonance Assignments ◽  
Human Immunoglobulin ◽  
Fc Fragment ◽  
Human Immunoglobulin G

scholarly journals Influence of Water Activity on Protease Adsorbed on Biochar in Organic Solvents

2017 ◽  
Vol 6 (4) ◽  
pp. 96 ◽  
Author(s):  
Hidetaka Noritomi ◽  
Jumpei Nishigami ◽  
Nobuyuki Endo ◽  
Satoru Kato ◽  
Katsumi Uchiyama
Keyword(s):  
Thermal Stability ◽  
Catalytic Activity ◽  
Organic Solvents ◽  
Water Activity ◽  
Initial Rate ◽  
Butyl Ester ◽  
Nitrogen Atmosphere ◽  
Bamboo Charcoal ◽  
Influence Of Water ◽  
Hydrolysis Of

We have found that the organic solvent-resistance of Alpha-chymotrypsin (Alpha-CT) is enhanced by adsorbing Alpha-CT onto bamboo charcoal powder (BCP), which is obtained by pyrolyzing bamboo waste under nitrogen atmosphere, and is markedly dependent on the thermodynamic water activity (aw) in organic solvents. When BCP-adsorbed　Alpha-CT was immersed in acetonitrile at an appropriate water activity, it effectively enhanced the transesterification of N-acetyl-L-tyrosine ethyl ester (N-Ac-Tyr-OEt) with n-butanol (BuOH) to produce N-acetyl-L-tyrosine butyl ester (N-Ac-Tyr-OBu), compared to the hydrolysis of N-Ac-Tyr-OEt with water to give N-acetyl-L-tyrosine (N-Ac-Tyr-OH). When the water activity was 0.28, the initial rate of transesterification catalyzed by BCP-adsorbed Alpha-CT was about sixty times greater than that catalyzed by free Alpha-CT. Regarding the reaction selectivity which is defined as a ratio of the initial rate of transesterification to that of hydrolysis, BCP-adsorbed α-CT was much superior to free Alpha-CT. The catalytic activity of BCP-adsorbed Alpha-CT was markedly dependent on the reaction temperature. Furthermore, concerning the thermal stability at 50 oC, the half-life of BCP-adsorbed Alpha-CT exhibited 3.8-fold, compared to that of free Alpha-CT.

scholarly journals Influence of Me2SO and incubation time on papain activity studied using fluorogenic substrates.

2001 ◽  
Vol 48 (4) ◽  
pp. 995-1002 ◽  
Author(s):  
M Szabelski ◽  
K Stachowiak ◽  
W Wiczk
Keyword(s):  
Incubation Time ◽  
Active Sites ◽  
Initial Rate ◽  
Substrate Binding ◽  
Rapid Decrease ◽  
Fluorogenic Substrates ◽  
Binding Process ◽  
Rate Of Hydrolysis ◽  
Hydrolysis Of

Papain activity in a buffer containing Me2SO was studied using fluorogenic substrates. It was found that the number of active sites of papain decreases with increasing Me2SO concentration whereas the incubation time, in a buffer containing 3% Me2SO does not affect the number of active sites. However, an increase of papain incubation time in the buffer with 3% Me2SO decreased the initial rate of hydrolysis of Z-Phe-Arg-Amc as well as Dabcyl-Lys-Phe-Gly-Gly-Ala-Ala-Edans. Moreover, an increase of Me2SO concentration in working buffer decreased the initial rate of papain-catalysed hydrolysis of both substrates. A rapid decrease of the initial rate (by up to 30%) was observed between 1 and 2% Me2SO. Application of the Michaelis-Menten equation revealed that at the higher Me2SO concentrations the apparent values of k(cat)/Km decreased as a result of Km increase and kcat decrease. However, Me2SO changed the substrate binding process more effectively (Km) than the rate of catalysis k(cat).

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