Amino Acid Residues His183 and Glu264 inBacillus subtilisFerrochelatase Direct and Facilitate the Insertion of Metal Ion into Protoporphyrin IX†,‡

Biochemistry ◽  
2007 ◽  
Vol 46 (1) ◽  
pp. 87-94 ◽  
Author(s):  
Mattias D. Hansson ◽  
Tobias Karlberg ◽  
Muhammad Arys Rahardja ◽  
Salam Al-Karadaghi ◽  
Mats Hansson
Keyword(s):  
Amino Acid ◽  
Metal Ion ◽  
Protoporphyrin Ix ◽  
Amino Acid Residues

scholarly journals The influence of sex steroid hormones on ferrochelatase gene expression in Harderian gland of hamster (Mesocricetus auratus)

2006 ◽  
Vol 189 (1) ◽  
pp. 103-112 ◽  
Author(s):  
F Vilchis ◽  
L Ramos ◽  
C Timossi ◽  
B Chávez
Keyword(s):  
Amino Acid ◽  
Steroid Hormones ◽  
Protoporphyrin Ix ◽  
Harderian Gland ◽  
Active Role ◽  
Structural Features ◽  
Sex Steroid Hormones ◽  
Sex Steroid ◽  
Amino Acid Residues ◽  
Haem Proteins

Ferrochelatase (protohaem ferrolyase, EC 4.99.1.1), the terminal enzyme of the haem biosynthetic pathway, catalyses the insertion of ferrous iron into protoporphyrin IX to form protohaem. The Syrian hamster Harderian gland (HG) is known for its ability to produce and accumulate large amounts of protoporphyrins. In this species, the female gland contains up to 120 times more porphyrin than the male gland. Data from biochemical studies suggest that this gland possesses the enzymatic complex for haem biosynthesis but lacks ferrochelatase activity. The abundance of intraglandular haem proteins does not support this idea. To gain more insight into this process, we isolated cDNA for ferrochelatase from hamster liver, using the 5′- and 3′- rapid amplification of complementary DNA ends (RACE), and investigated its expression in HG from males and females. The full-length cDNA comprises an open reading frame of 1269 bp encoding a polypeptide of 422 amino-acid residues. Hamster DNA sequence exhibits 92% identity to mouse and 87% identity to human sequences. The predicted hamster enzyme was shown to have structural features of mammalian ferrochelatase, including a putative NH2- terminal presequence, a central core of about 330 amino-acid residues and an extra 30–50-amino-acid stretch at the carboxyl-terminus. RNA blotting experiments indicated that this cDNA hybridized to a liver mRNA of about 2.1 kb, while a weak hybridization signal was observed with mRNA from HG preparations. RT–PCR assays confirmed the expression of specific transcripts in both tissues. Male glands contained approximately twofold more enzyme mRNA than female glands. Likewise, the intraglandular content of mRNA varied during the oestrous cycle, with the highest levels found in the oestrous phase. These cyclic variations were less evident in liver. Ovariectomy plus treatment with progesterone or 17β-oestradiol plus progesterone increased ferrochelatase mRNA of the gland. In HG of short- or long-term castrated males, the administration of testosterone did not affect the ferrochelatase mRNA concentration. Based on mRNA expression levels, we conclude that Harderian ferrochelatase may play an active role in maintaining the physiological pool of haem required for processing cytochromes and other glandular haem proteins. Likewise, the sex-steroid hormones appear to have only a modest influence upon Harderian ferrochelatase.

scholarly journals Decoding Essential Amino Acid Residues in the Substrate Groove of a Non-Specific Nuclease from Pseudomonas syringae

Catalysts ◽  
2019 ◽  
Vol 9 (11) ◽  
pp. 941 ◽  
Author(s):  
Lynn Sophie Schwardmann ◽  
Sarah Schmitz ◽  
Volker Nölle ◽  
Skander Elleuche
Keyword(s):  
Amino Acid ◽  
Pseudomonas Syringae ◽  
Metal Ion ◽  
Homology Model ◽  
Downstream Processing ◽  
Outer Ring ◽  
Amino Acid Residues ◽  
Independent Manner ◽  
Identify Amino Acid ◽  
Close Proximity

Non-specific nucleases (NSN) are of interest for biotechnological applications, including industrial downstream processing of crude protein extracts or cell-sorting approaches in microfabricated channels. Bacterial nucleases belonging to the superfamily of phospholipase D (PLD) are featured for their ability to catalyze the hydrolysis of nucleic acids in a metal-ion-independent manner. In order to gain a deeper insight into the composition of the substrate groove of a NSN from Pseudomonas syringae, semi-rational mutagenesis based on a structure homology model was applied to identify amino acid residues on the protein’s surface adjacent to the catalytic region. A collection of 12 mutant enzymes each with a substitution to a positively charged amino acid (arginine or lysine) was produced in recombinant form and biochemically characterized. Mutations in close proximity to the catalytic region (inner ring) either dramatically impaired or completely abolished the enzymatic performance, while amino acid residues located at the border of the substrate groove (outer ring) only had limited or no effects. A K119R substitution mutant displayed a relative turnover rate of 112% compared to the original nuclease. In conclusion, the well-defined outer ring of the substrate groove is a potential target for modulation of the enzymatic performance of NSNs belonging to the PLD superfamily.

Design of Novel Hemoglobins

1997 ◽  
Author(s):  
C. Ho ◽  
H.-W. Kim
Keyword(s):  
Amino Acid ◽  
Oxygen Carrier ◽  
Protoporphyrin Ix ◽  
Iron Complex ◽  
Heme Iron ◽  
Oxygen Binding ◽  
Amino Acid Residues ◽  
Allosteric Proteins ◽  
Cooperative Process ◽  
Mutant Forms

Human normal adult hemoglobin (Hb) A, the oxygen carrier of blood, is a tetrameric protein consisting of two α chains of 141 amino acid residues each and two β chains of 146 amino acid residues each. Each Hb chain contains a heme group which is an iron complex of protoporphyrin IX. Under physiological conditions, the heme-iron atoms of Hb remain in the ferrous state. In the absence of oxygen, the four heme-irom atoms in Hb A are in the highspin ferrous state [Fe(II)] with four unpaired electrons each. Each of the four heme-iron atoms in Hb A can combine with an O2 molecule to give oxyhemoglobin (HbO2) in which the iron atom is in a low-spin, diamagnetic ferrous state. The oxygen binding of Hb exhibits sigmoidal behavior, with an overall association constant expression giving a greater than first-power dependence on the concentration of O2. Thus, the oxygenation of Hb is a cooperative process, such that when one O2 is bound, succeeding O2 molecules are bound more readily. Hb is an allosteric protein, i.e., its functional properties are regulated by a number of metabolites [such as hydrogen ions, chloride, carbon dioxide, 2,3-diphosphoglycerate (2,3-DPG)] other than its ligand, O2. It has been used as a model for allosteric proteins, and indeed, hemoglobins of vertebrates are among the most extensively studied allosteric proteins. Their allosteric properties are physiologically important in optimizing O2 transport by erythrocytes. The large number of mutant forms of Hb available provides an array of structural alterations with which to correlate effects on function. For details, see DickersonandGeis (1983), Bunnand Forget (1986), Ho (1992), Ho and Perussi (1994). There are two types of contacts between the α and β subunits of Hb (Perutz, 1970; Dickerson and Geis, 1983). The α1β1 (or α2 β2) contacts, involving B, G, and H helices, and GH corners, are called packing contacts. These contacts remain unchanged and hold the dimer together even when there is a change in the ligation state of the heme.

Acid-base and metal ion complex formation properties of polymers containing amino acid residues

1986 ◽  
Vol 19 (1) ◽  
pp. 37-42 ◽  
Author(s):  
Rolando Barbucci ◽  
Mario Casolaro ◽  
Mila Nocentini ◽  
Silvia Corezzi ◽  
Paolo Ferruti ◽  
...  
Keyword(s):  
Amino Acid ◽  
Complex Formation ◽  
Metal Ion ◽  
Amino Acid Residues ◽  
Acid Base ◽  
Polymers Containing Amino Acid ◽  
Metal Ion Complex

scholarly journals Purification, Characterization, and Gene Cloning of Purine Nucleosidase from Ochrobactrum anthropi

2001 ◽  
Vol 67 (4) ◽  
pp. 1783-1787 ◽  
Author(s):  
Jun Ogawa ◽  
Sou Takeda ◽  
Sheng-Xue Xie ◽  
Haruyo Hatanaka ◽  
Toshihiko Ashikari ◽  
...  
Keyword(s):  
Amino Acid ◽  
Metal Ion ◽  
Amino Acid Sequences ◽  
Leader Peptide ◽  
Amino Acid Residues ◽  
Ochrobactrum Anthropi ◽  
Pyrimidine Nucleotides ◽  
Reading Frame ◽  
Pyrimidine Nucleosides ◽  
Nicotinamide Mononucleotide

ABSTRACT A bacterium, Ochrobactrum anthropi, produced a large amount of a nucleosidase when cultivated with purine nucleosides. The nucleosidase was purified to homogeneity. The enzyme has a molecular weight of about 170,000 and consists of four identical subunits. It specifically catalyzes the irreversibleN-riboside hydrolysis of purine nucleosides, theKm values being 11.8 to 56.3 μM. The optimal activity temperature and pH were 50°C and pH 4.5 to 6.5, respectively. Pyrimidine nucleosides, purine and pyrimidine nucleotides, NAD, NADP, and nicotinamide mononucleotide are not hydrolyzed by the enzyme. The purine nucleoside hydrolyzing activity of the enzyme was inhibited (mixed inhibition) by pyrimidine nucleosides, with Ki and Ki ′ values of 0.455 to 11.2 μM. Metal ion chelators inhibited activity, and the addition of Zn2+ or Co2+ restored activity. A 1.5-kb DNA fragment, which contains the open reading frame encoding the nucleosidase, was cloned, sequenced, and expressed inEscherichia coli. The deduced 363-amino-acid sequence including a 22-residue leader peptide is in agreement with the enzyme molecular mass and the amino acid sequences of NH2-terminal and internal peptides, and the enzyme is homologous to known nucleosidases from protozoan parasites. The amino acid residues forming the catalytic site and involved in binding with metal ions are well conserved in these nucleosidases.

α-Glutamic acid-β-naphthylamidase from Neisseria catarrhalis

1969 ◽  
Vol 15 (11) ◽  
pp. 1293-1300 ◽  
Author(s):  
Sidney T. Cox ◽  
Francis J. Behal
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Glutamic Acid ◽  
Metal Ion ◽  
Gel Filtration ◽  
Aminopeptidase Activity ◽  
Amino Acid Residues ◽  
Ph Optimum ◽  
Hydrolysis Of ◽  
Derivatives Of

A second bacterial peptidase-like enzyme, arylamidase-II, has been isolated from cell free extracts of Neisseria catarrhalis. Arylamidase-II action is limited primarily to the hydrolysis of α-glutamic acid and α-aspartic acid derivatives of β-naphthylamine and short peptides of glutamic acid. The enzyme was purified 450-fold by gel filtration, ion exchange, and calcium phosphate chromatography. Its pH optimum and molecular weight were 7.7 and 170 000, respectively. Aside from its restricted substrate specificity, arylamidase-II has been found to be closely related in its mechanism of action, molecular weight, pH optimum, and metal ion dependence to arylamidase-I, which catalyzes the hydrolysis of neutral amino acid derivatives of β-naphthylamine. Arylamidase-II exhibits aminopeptidase activity, requiring the amino acid residues in the N-terminal and penultimate position to be of the L-configuration in order for hydrolysis to occur.

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