Structure and Proposed Amino-Acid Sequence of a Pepsin from Atlantic Cod (Gadus morhua)

1998 ◽  
Vol 54 (1) ◽  
pp. 32-46 ◽  
Author(s):  
Solveig Karlsen ◽  
Edward Hough ◽  
Ragnar L. Olsen
Keyword(s):  
Crystal Structure ◽  
Amino Acid ◽  
Gadus Morhua ◽  
Model Building ◽  
Atlantic Cod ◽  
Tuna Fish ◽  
Area Detector ◽  
Replacement Method ◽  
Molecular Replacement Method ◽  
Starting Model

The crystal structure of a pepsin from the gastric mucosa of Atlantic cod has been determined to 2.16 Å resolution. Data were collected on orthorhombic crystals with cell dimensions a = 35.98, b = 75.40 and c = 108.10 Å, on a FAST area-detector system. The phase problem was solved by the molecular-replacement method using porcine pepsin (PDB entry 5PEP) as a search model. The structure has been refined to a crystallographic R factor of 20.8% using all reflections between 8.0 and 2.16 Å, without prior knowedge of the primary sequence. The resulting crystal structure is very similar to the porcine enzyme, consisting of two domains with predominantly β-sheet structure in the same sequential positions as the enzyme from pig. In the course of the model building, 122 residues were substituted and two residues deleted from the starting model to give a polypeptide chain of 324 amino acids and a sequence identity of 57.7% with the pig pepsin. No carbohydrate residues were located. Sequence alignment with available aspartic proteinases, indicates that the fish enzyme seems to be more related to mammalian gastric pepsins than to the mammalian gastricsins and chymosins, lysosomal cathepsin D's and a pepsin from tuna fish. The amino-acid composition of the cod enzyme, however, is more in accordance with the cathepsin D's.

scholarly journals Crystal structure of the aromatic-amino-acid aminotransferase fromStreptococcus mutans

2019 ◽  
Vol 75 (2) ◽  
pp. 141-146
Author(s):  
Xuzhen Cong ◽  
Xiaolu Li ◽  
Shentao Li
Keyword(s):  
Crystal Structure ◽  
Amino Acid ◽  
Streptococcus Mutans ◽  
Aromatic Amino Acid ◽  
Aromatic Amino Acids ◽  
Diffusion Method ◽  
Molecular Replacement ◽  
Replacement Method ◽  
Molecular Replacement Method ◽  
Aromatic Amino Acid Aminotransferase

Streptococcus mutans, a facultatively aerobic and Gram-positive bacterium, is the primary causative agent of dental caries and contributes to the multispecies biofilm known as dental plaque. In this study, the aromatic-amino-acid aminotransferase fromStreptococcus mutans(SmAroAT) was recombinantly expressed inEscherichia coli. An effective purification protocol was established. The recombinant protein was crystallized using the hanging-drop vapor-diffusion method with PEG 3350 as the primary precipitant. The crystal structure ofSmAroAT was solved at 2.2 Å resolution by the molecular-replacement method. Structural analysis indicated that the proteins of the aromatic-amino-acid aminotransferase family have conserved structural elements that might play a role in substrate binding. These results may help in obtaining a better understanding of the catabolism and biosynthesis of aromatic amino acids.

Effect of age and temperature on amino acid composition and the content of different protein types of juvenile Atlantic cod (Gadus morhua) otoliths

2004 ◽  
Vol 61 (6) ◽  
pp. 1012-1020 ◽  
Author(s):  
K Hüssy ◽  
H Mosegaard ◽  
F Jessen
Keyword(s):  
Amino Acid ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Soluble Protein ◽  
Protein Fraction ◽  
Gadus Morhua ◽  
Atlantic Cod ◽  
Water Soluble ◽  
Soluble Protein Fraction ◽  
Water Soluble Protein

The purpose of this study was to analyse the amino acid composition of otolith matrix protein, estimate the proportion of the water-soluble protein fraction, and analyse the effect of matrix composition on otolith visual appearance. Juvenile Atlantic cod (Gadus morhua) were reared under constant temperature and feeding conditions and sampled at the beginning and the end of the experiment. The amino acid composition was dominated by asparagine, glutamic acid, leucine, serine, and proline. A change in amino acid composition was observed with increasing temperature and time, caused by changing proportions of the water-soluble and -insoluble protein fractions. Feeding level had no effect. The relative content of water-soluble protein was linearly related to fish dry weight and temperature. Otolith opacity, defined as the percentage of incident light absorbed by an otolith section, did not differ significantly between experimental treatments. The soluble protein fraction had a positive, albeit insignificant, correlation with opacity. Using opacity and otolith volume, deposited total otolith protein content was estimated with an R2 of 0.91, where otolith volume alone explained 83% of the observed variation.

Crystallization and structure solution of p53 (residues 326–356) by molecular replacement using an NMR model as template

1998 ◽  
Vol 54 (1) ◽  
pp. 86-89 ◽  
Author(s):  
Peer R. E. Mittl ◽  
Patrick Chène ◽  
Markus G. Grütter
Keyword(s):  
Crystal Structure ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Molecular Replacement ◽  
Additional Information ◽  
Nmr Structures ◽  
Replacement Method ◽  
Tetramerization Domain ◽  
Structure Solution ◽  
Molecular Replacement Method

The molecular replacement method is a powerful technique for crystal structure solution but the use of NMR structures as templates often causes problems. In this work the NMR structure of the p53 tetramerization domain has been used to solve the crystal structure by molecular replacement. Since the rotation- and translation-functions were not sufficiently clear, additional information about the symmetry of the crystal and the protein complex was used to identify correct solutions. The three-dimensional structure of residues 326–356 was subsequently refined to a final R factor of 19.1% at 1.5 Å resolution.

scholarly journals Nucleotide Polymorphism and Natural Selection at the Pantophysin (Pan I) Locus in the Atlantic Cod, Gadus morhua (L.)

Genetics ◽  
2001 ◽  
Vol 157 (1) ◽  
pp. 317-330 ◽  
Author(s):  
Grant H Pogson
Keyword(s):  
Natural Selection ◽  
Amino Acid ◽  
Gadus Morhua ◽  
Atlantic Cod ◽  
Amino Acid Level ◽  
Amino Acid Substitutions ◽  
Significant Heterogeneity ◽  
Nucleotide Sequence Variation ◽  
Spatially Varying ◽  
Spatially Varying Selection

Abstract Molecular studies of nucleotide sequence variation have rarely attempted to test hypotheses related to geographically varying patterns of natural selection. The present study tested the role of spatially varying selection in producing significant linkage disequilibrium and large differences in the frequencies of two common alleles at the pantophysin (Pan I) locus among five populations of the Atlantic cod, Gadus morhua. Nucleotide sequences of 124 Pan I alleles showed strong evidence for an unusual mix of balancing and directional selection but no evidence of stable geographically varying selection. The alleles were highly divergent at both the nucleotide level (differing on average by 19 mutations) and at amino acid level (each having experienced three amino acid substitutions since diverging from a common ancestral allele). All six amino acid substitutions occurred in a 56-residue intravesicular loop (IV1 domain) of the vesicle protein and each involved a radical change. An analysis of molecular variation revealed significant heterogeneity in the frequencies of recently derived mutations segregating within both allelic classes, suggesting that two selective sweeps may be presently occurring among populations. The dynamic nature of the Pan I polymorphism in G. morhua and clear departure from equilibrium conditions invalidate a simple model of spatially varying selection.

Antifreeze glycoproteins in the plasma of Newfoundland Atlantic cod (Gadus morhua)

1981 ◽  
Vol 59 (11) ◽  
pp. 2186-2192 ◽  
Author(s):  
Choy L. Hew ◽  
Don Slaughter ◽  
Garth L. Fletcher ◽  
Shashikant B. Joshi
Keyword(s):  
Amino Acid ◽  
High Performance ◽  
Gadus Morhua ◽  
Atlantic Cod ◽  
Antifreeze Proteins ◽  
Gel Filtration ◽  
Exchange Chromatography ◽  
Polar Cod ◽  
Antifreeze Glycoproteins ◽  
Molecular Weights

The plasma of the Atlantic cod, Gadus morhua, contained antifreeze glycoproteins which were present only during the winter months. The antifreeze proteins were isolated, using gel filtration and ion exchange chromatography, and characterized by high performance liquid chromatography. The antifreeze proteins appeared to consist of at least seven components with molecular weights ranging from 2 500 to 33 000. Chemical analysis of the larger components showed a predominance of alanine, threonine, and galactosamine in a ratio of 2:1:1. The smaller peptides contained proline, in addition to alanine and threonine. The amino acid sequence of the smallest glycopeptide (molecular weight 2500) was found to be Ala Ala Thr Pro Ala Thr Ala Ala Thr Pro Ala Thr Ala Ala.These glycoproteins are very similar, if not identical, in amino acid and carbohydrate composition to those isolated from Antaractic nototheniids and several northern gadoids. The sequence of the smallest glycopeptide from the Atlantic cod is identical to that reported for the polar cod, Boreogadus saida.

Effects of amino acid supplementations on metabolic and physiological parameters in Atlantic cod (Gadus morhua) under stress

2016 ◽  
Vol 43 (2) ◽  
pp. 591-602 ◽  
Author(s):  
Marcelino Herrera ◽  
María Antonia Herves ◽  
Inmaculada Giráldez ◽  
Kristin Skar ◽  
Hanne Mogren ◽  
...  
Keyword(s):  
Amino Acid ◽  
Gadus Morhua ◽  
Atlantic Cod ◽  
Physiological Parameters

scholarly journals Crystal Structure of BRP39 protein expressed during Mammary Gland Apoptosis

2014 ◽  
Vol 70 (a1) ◽  
pp. C1638-C1638
Author(s):  
Suman Choudhary ◽  
Ashok Mohanty ◽  
Andrew Fisher
Keyword(s):  
Breast Cancer ◽  
Crystal Structure ◽  
Carbohydrate Binding ◽  
Potential Candidate ◽  
Structure Based Drug Design ◽  
Sugar Binding ◽  
Replacement Method ◽  
Tim Barrel ◽  
Molecular Replacement Method ◽  
Binding Groove

Breast regression protein (BRP39) from mice is a 39 kDa protein which is a member of chitolectin class of GH18 family. High levels of expression of BRP39 have been detected in breast carcinoma. It has been proposed that this may act as a protecting signalling factor and also help in proliferation of cells during breast cancer. It can act as a potential candidate for rational structure based drug design against breast cancer. Here, we report the crystal structure of recombinant BRP39 in a deglycosylated form which was expressed in a heterologous system i.e E.Coli. To understand the role of sugar moiety, crystal structure of a deglycosylated chitolectin is an essential requirement. The structure was solved by molecular replacement method of phase determination and refined to a 2.6 A0resolution. The overall structure of BRP39 consists of two globular domains: a large (β/α)8 TIM barrel domain and a small (α+β) domain. The most striking observation from BRP39 structure is the conformation of critical Trp 100 residue into the β-barrel of carbohydrate binding groove of BRP39. The structure reveals that the glycan moiety plays an important role in the orientation of critical Trp100 in the β-barrel. In deglycosylated BRP39, it orients away from the barrel and resembles the conformation as seen in non-glycosylated chitinases. In contrast to this, the corresponding Trp is oriented into the barrel in case of other glycosylated homologues of BRP39 which may have its implications in sugar binding. Furthermore, in MGP-40, the altered conformation of loop is stabilized by H-bonding and stacking interactions whereas in BRP39, no hydrogen bond was observed between any of the residues of this loop except for Trp100 which interacts with a water molecule may be to stabilize its conformation. Another important observation in the sugar binding groove of BRP39 structure is the mutation of two important residues, one in TIM barrel domain and another in a part of α+β domain which is also involved in sugar binding. Asn100 and Arg263 in Hcgp39 and other chitinase- like proteins (SPX-40 structures) are replaced by Lys101 and Lys264 in BRP39. Both of these residues i.e. Asn100 and Arg 263 in HCgp39 have been reported to be involved in stabilizing the binding of GlcNAc inside the groove. Possibly, there is a significant distortion in the shape of sugar binding groove due to these mutations in BRP39.

Amino Acid Decarboxylase Activity and Other Chemical Characteristics as Related to Freshness Loss in Iced Cod (Gadus morhua)†

2002 ◽  
Vol 65 (7) ◽  
pp. 1152-1157 ◽  
Author(s):  
M. MANUELA HERNÁNDEZ-HERRERO ◽  
GUILLAUME DUFLOS ◽  
PIERRE MALLE ◽  
STÉPHANE BOUQUELET
Keyword(s):  
Amino Acid ◽  
Biogenic Amine ◽  
Gadus Morhua ◽  
Atlantic Cod ◽  
Storage Period ◽  
Acid Decarboxylase ◽  
Decarboxylase Activity ◽  
Exponential Relationship ◽  
Sensory Score ◽  
Iced Storage

Biogenic amine levels and other biochemical indicators were measured to study the safety of and the loss of freshness in iced Atlantic cod. Biogenic amine content exhibited high variability during iced storage of Atlantic cod. Ornithine and lysine decarboxylase activity apparently increased at the end of the storage period. Amino acid activity was probably generated by endogenous amino acid decarboxylases of raw fish. No statistical differences were observed in the total volatile base fraction or in the ammonia or monomethylamine contents during iced storage. However, trimethylamine contents showed a significant exponential relationship with time and sensory score. Cod formed inosine as the major metabolite of IMP. The H and G indices showed a linear relationship with time and sensory score and served as good indicators of cod freshness quality. However, the K, Ki, and P indices showed a logarithmic relationship with time and sensory score. IMP, K, Ki, and P served as indicators of freshness lost during the early stages of chilled storage of cod.

scholarly journals Sequence Analysis and Preliminary X-ray Crystallographic Analysis of an Acetylesterase (LgEstI) from Lactococcus garvieae

Crystals ◽  
2021 ◽  
Vol 12 (1) ◽  
pp. 46
Author(s):  
Hackwon Do ◽  
Ying Wang ◽  
Chang Woo Lee ◽  
Wanki Yoo ◽  
Sangeun Jeon ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Protein Engineering ◽  
Model Building ◽  
Structure Refinement ◽  
Crystallographic Analysis ◽  
Wild Type ◽  
Acetyl Esterase ◽  
X Ray ◽  
Lactococcus Garvieae ◽  
Replacement Method

A gene encoding LgEstI was cloned from a bacterial fish pathogen, Lactococcus garvieae. Sequence and bioinformatic analysis revealed that LgEstI is close to the acetyl esterase family and had maximum similarity to a hydrolase (UniProt: Q5UQ83) from Acanthamoeba polyphaga mimivirus (APMV). Here, we present the results of LgEstI overexpression and purification, and its preliminary X-ray crystallographic analysis. The wild-type LgEstI protein was overexpressed in Escherichia coli, and its enzymatic activity was tested using p-nitrophenyl of varying lengths. LgEstI protein exhibited higher esterase activity toward p-nitrophenyl acetate. To better understand the mechanism underlying LgEstI activity and subject it to protein engineering, we determined the high-resolution crystal structure of LgEstI. First, the wild-type LgEstI protein was crystallized in 0.1 M Tris-HCl buffer (pH 7.1), 0.2 M calcium acetate hydrate, and 19% (w/v) PEG 3000, and the native X-ray diffraction dataset was collected up to 2.0 Å resolution. The crystal structure was successfully determined using a molecular replacement method, and structure refinement and model building are underway. The upcoming complete structural information of LgEstI may elucidate the substrate-binding mechanism and provide novel strategies for subjecting LgEstI to protein engineering.

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