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1.
Because free amino acids were most probably available in primitive abiotic environments, their metabolism is likely to have provided some of the very first metabolic pathways of life. What were the first enzymatic reactions to emerge? A cladistic analysis of metabolic pathways of the 16 aliphatic amino acids and 2 portions of the Krebs cycle was performed using four criteria of homology. The analysis is not based on sequence comparisons but, rather, on coding similarities in enzyme properties. The properties used are shared specific enzymatic activity, shared enzymatic function without substrate specificity, shared coenzymes, and shared functional family. The tree shows that the earliest pathways to emerge are not portions of the Krebs cycle but metabolisms of aspartate, asparagine, glutamate, and glutamine. The views of Horowitz (Horowitz, N. H. (1945) Proc. Natl. Acad. Sci. U. S. A. 31, 153-157) and Cordón (Cordón, F. (1990) Tratado Evolucionista de Biologia, Aguilar, Madrid, Spain), according to which the upstream reactions in the catabolic pathways and the downstream reactions in the anabolic pathways are the earliest in evolution, are globally corroborated; however, with some exceptions. These are due to later opportunistic connections of pathways (actually already suggested by these authors). Earliest enzymatic functions are mostly catabolic; they were deaminations, transaminations, and decarboxylations. From the consensus tree we extracted four time spans for amino acid metabolism development. For some amino acids catabolism and biosynthesis occurred at the same time (Asp, Glu, Lys, Leu, Ala, Val, Ile, Pro, Arg). For others ultimate reactions that use amino acids as a substrate or as a product are distinct in time, with catabolism preceding anabolism for Asn, Gln, and Cys and anabolism preceding catabolism for Ser, Met, and Thr. Cladistic analysis of the structure of biochemical pathways makes hypotheses in biochemical evolution explicit and parsimonious. 相似文献
2.
Metabolic pathways exhibit structures resulting from an evolutionary process. Pathways have been inherited through time with modification, from the earliest periods of life. It is possible to compare the structure of pathways as done in comparative anatomy, i.e. for inferring ancestral pathways or parts of it (ancestral enzymatic functions), using standard phylogenetic reconstruction. Thus a phylogenetic tree of pathways provides a relative ordering of the rise of enzymatic functions. It even becomes possible to order the birth of each complete pathway in time. This particular "DNA-free" conceptual approach to evolutionary biochemistry is reviewed, gathering all the justifications given for it. Then, the method of assigning a given pathway to a time span of biochemical development is revisited. The previous method used an implicit "clock" of metabolic development that is difficult to justify. We develop a new clock-free approach, using functional biochemical arguments. Results of the two methods are not significantly different; our method is just more precise. This suggests that the clock assumed in the first method does not provoke any important artefact in describing the development of biochemical evolution. It is just unnecessary to postulate it. As a result, most of the amino acid metabolic pathways develop forwards, confirming former models of amino acid catabolism evolution, but not those for amino acid anabolism. The order of appearance of sectors of universal cellular metabolism is: (1) amino acid catabolism, (2) amino acid anabolism and closure of the urea cycle, (3) glycolysis and glycogenesis, (4) closure of the pentose-phosphate cycle, (5) closure of the Krebs cycle and fatty acids metabolism, (6) closure of the Calvin cycle. 相似文献
3.
The acidic amino acids (Asp, Glu) and their amides (Asn, Gln) support rapid growth of a variety of Pseudomonas strains when provided as the sole source of carbon and nitrogen. All key enzymes of glutamate metabolism were detected in P. fluorescence, with glutaminase and asparaginase showing the highest specific activities. A periplasmic glutaminase/asparaginase activity (PGA) was found in all pseudomonads examined, including a number of root-colonizing biocontrol strains. The enzyme was purified and shown to be identical with the ansB gene product described previously. In addition to PGA, P. fluorescens contains a cytoplasmic asparaginase with marked specificity for Asn. PGA is strongly and specifically induced by its substrates (Asn, Gln) but also by the reaction products (Asp, Glu). In addition, PGA is subject to efficient carbon catabolite repression by glucose and by citrate cycle metabolites. A mutant of P. putida KT2440 with a disrupted ansB gene was unable to utilize Gln, whereas growth of the mutant on other amino acids was normal. 相似文献
4.
Antonio H. Lancha Jr Jacques R. Poortmans Luciana O. Pereira 《Cell biochemistry and function》2009,27(8):552-557
The consumption of protein supplements containing amino acids is increasing around the world. Aspartate (Asp) and asparagine (Asn) are amino acids metabolized by skeletal muscle. This metabolism involves biochemical pathways that are involved in increasing Krebs cycle activity via anaplerotic reactions, resulting in higher glutamine concentrations. A connection between amino acid supplementation, glycogen concentration, and glucose uptake has been previously demonstrated. The purpose of this study was to evaluate the effect of Asp and Asn supplementation on glucose uptake in rats using three different glycogen concentrations. The results indicate that Asp and Asn supplementation in rats with high glycogen concentrations (fed state) further increased the glycogen concentration in the muscle, and decreased in vitro 2‐deoxyglucose (a glucose analog) uptake by the muscle at maximal insulin concentrations. When animals had a medium glycogen concentration (consumed lard for 3 days), glucose uptake was higher in the supplemented group at sub‐maximal insulin concentrations. We conclude that supplementation of Asp and Asn reduced glucose transport in rat muscle only at higher levels of glycogen. The ingestion of lard for 3 days changed the responsiveness and sensitivity to insulin, and that group had higher levels of insulin sensitivity with Asp and Asn supplementation. Copyright © 2009 John Wiley & Sons, Ltd. 相似文献
5.
The mammalian Na(+)/H(+) exchanger isoform 1 (NHE1) resides on the plasma membrane and exchanges one intracellular H(+) for one extracellular Na(+). It maintains intracellular pH and regulates cell volume, and cell functions including growth and cell differentiation. Previous structural and functional studies on TMVI revealed several amino acids that are potentially pore lining. We examined these and other critical residues by site-directed mutagenesis substituting Asn227→Ala, Asp, Arg; Ile233→Ala; Leu243→Ala; Glu247→Asp, Gln; Glu248→Asp, Gln. Mutant NHE1 proteins were characterized in AP-1 cells, which do not express endogenous NHE1. All the TMVI critical amino acids were highly sensitive to substitution and changes often lead to a dysfunctional protein. Mutations of Asn227→Ala, Asp, Arg; Ile233→Ala; Leu243→Ala; Glu247→Asp; Glu248→Gln yielded significant reduction in NHE1 activity. Mutants of Asn227 demonstrated defects in protein expression, targeting and activity. Substituting Asn227→Arg and Ile233→Ala decreased the surface localization and expression of NHE1 respectively. The pore lining amino acids Ile233 and Leu243 were both essential for activity. Glu247 was not essential, but the size of the residue at this location was important while the charge on residue Glu248 was more critical to NHE1 function. Limited trypsin digestion on Leu243→Ala and Glu248→Gln revealed that they had increased susceptibility to proteolytic attack, indicating an alteration in protein conformation. Modeling of TMVI with TMXI suggests that these TM segments form part of the critical fold of NHE1 with Ile233 and Leu465 of TMXI forming a critical part of the extracellular facing ion conductance pathway. 相似文献
6.
15N-labelled (amino group) asparagine (Asn), glutamate (Glu), alanine (Ala), aspartate (Asp) and serine (Ser) were used to study the metabolic role and the participation of each compound in the photorespiratory N cycle ofPisum sativum L. leaves. Asparagine was utilised as a nitrogen source by either deamidation or transamination, Glu was converted to Gln through NH3 assimilation and was a major amino donor for transamination, and Ala was utilised by transamination to a range of amino acids. Transamination also provided a pathway for Asp utilisation, although Asp was also used as a substrate for Asn synthesis. In the photorespiratory synthesis of glycine (Gly), Ser, Ala, Glu and Asn acted as sources of amino-N, contributing, in the order given, 38, 28, 23, and 7% of the N for glycine synthesis; Asp provided less than 4% of the amino-N in glycine. Calculations based on the incorporation of15N into Gly indicated that about 60% (Ser), 20% (Ala), 12% (Glu) and 11% (Asn) of the N metabolised from each amino acid was utilised in the photorespiratory nitrogen cycle.Abbreviations Ala
alamine
- Asn
asparagine
- Asp
aspartate
- Glu
glutamate
- MOA
methoxylamine
- Ser
serine 相似文献
7.
Roles of glutamine in neurotransmission 总被引:1,自引:0,他引:1
Glutamine (Gln) is found abundantly in the central nervous system (CNS) where it participates in a variety of metabolic pathways. Its major role in the brain is that of a precursor of the neurotransmitter amino acids: the excitatory amino acids, glutamate (Glu) and aspartate (Asp), and the inhibitory amino acid, γ-amino butyric acid (GABA). The precursor-product relationship between Gln and Glu/GABA in the brain relates to the intercellular compartmentalization of the Gln/Glu(GABA) cycle (GGC). Gln is synthesized from Glu and ammonia in astrocytes, in a reaction catalyzed by Gln synthetase (GS), which, in the CNS, is almost exclusively located in astrocytes (Martinez-Hernandez et al., 1977). Newly synthesized Gln is transferred to neurons and hydrolyzed by phosphate-activated glutaminase (PAG) to give rise to Glu, a portion of which may be decarboxylated to GABA or transaminated to Asp. There is a rich body of evidence which indicates that a significant proportion of the Glu, Asp and GABA derived from Gln feed the synaptic, neurotransmitter pools of the amino acids. Depolarization-induced-, calcium- and PAG activity-dependent releases of Gln-derived Glu, GABA and Asp have been observed in CNS preparations in vitro and in the brain in situ. Immunocytochemical studies in brain slices have documented Gln transfer from astrocytes to neurons as well as the location of Gln-derived Glu, GABA and Asp in the synaptic terminals. Patch-clamp studies in brain slices and astrocyte/neuron co-cultures have provided functional evidence that uninterrupted Gln synthesis in astrocytes and its transport to neurons, as mediated by specific carriers, promotes glutamatergic and GABA-ergic transmission. Gln entry into the neuronal compartment is facilitated by its abundance in the extracellular spaces relative to other amino acids. Gln also appears to affect neurotransmission directly by interacting with the NMDA class of Glu receptors. Transmission may also be modulated by alterations in cell membrane polarity related to the electrogenic nature of Gln transport or to uncoupled ion conductances in the neuronal or glial cell membranes elicited by Gln transporters. In addition, Gln appears to modulate the synthesis of the gaseous messenger, nitric oxide (NO), by controlling the supply to the cells of its precursor, arginine. Disturbances of Gln metabolism and/or transport contribute to changes in Glu-ergic or GABA-ergic transmission associated with different pathological conditions of the brain, which are best recognized in epilepsy, hepatic encephalopathy and manganese encephalopathy. 相似文献
8.
This is report of mutational analysis of higher plant 5'-methylthioadenosine nucleosidase (MTAN). We identified and characterized the gene encoding yellow lupine (Lupinus luteus) MTAN (LlMTAN). The role of active site amino acids residues Glu24, Phe134, Glu188 and Asp211 was analyzed by site-directed mutagenesis. The Glu24Gln and Asp211Asn substitutions completely abolished the enzyme activity. The Glu188Gln mutant showed only trace activity toward 5'-methylthioadenosine. These results indicate that these three amino acid residues are necessary for enzyme activity. Furthermore, as the result of replacement of Phe134 by less bulky leucine, LlMTAN acquired the ability to bind and hydrolyze S-adenosylhomocysteine. We also analyzed the sequence of the LlMTAN promoter region. It appeared that there may be a direct link between LlMTAN expression regulation and sulfate metabolism. 相似文献
9.
Mutation of active site residues in synthetic T4-lysozyme gene and their effect on lytic activity 总被引:1,自引:0,他引:1
N N Anand E R Stephen S A Narang 《Biochemical and biophysical research communications》1988,153(2):862-868
The active site amino acids (Glu11 and Asp20) in T4-lysozyme have been mutated to their isosteric residues Gln or Asn and/or acidic residues such as Glu----Asp or Asp----Glu by the oligonucleotide-replacement method. Out of eight mutants so generated the mutant T4-lysozyme obtained from pTLY.Asp11 retains maximum amount of activity (approximately 16%), pTLY.Asn20 the least (0.9%) whereas pTLY.Gln11 lost completely. A systematic study of the active and inactive mutants thus generated supports the important role of Glu11 and Asp20 in T4-lysozyme activity as predicted in earlier studies. 相似文献
10.
Increased lysine synthesis coupled with a knockout of its catabolism synergistically boosts lysine content and also transregulates the metabolism of other amino acids in Arabidopsis seeds 总被引:1,自引:0,他引:1
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To elucidate the relative significance of Lys synthesis and catabolism in determining Lys level in plant seeds, we expressed a bacterial feedback-insensitive dihydrodipicolinate synthase (DHPS) in a seed-specific manner in wild-type Arabidopsis as well as in an Arabidopsis knockout mutant in the Lys catabolism pathway. Transgenic plants expressing the bacterial DHPS, or the knockout mutant, contained approximately 12-fold or approximately 5-fold higher levels, respectively, of seed free Lys than wild-type plants. However, the combination of these two traits caused a synergistic approximately 80-fold increase in seed free Lys level. The dramatic increase in free Lys in the knockout mutant expressing the bacterial DHPS was associated with a significant reduction in the levels of Glu and Asp but also with an unexpected increase in the levels of Gln and Asn. This finding suggested a special regulatory interaction between Lys metabolism and amide amino acid metabolism in seeds. Notably, the level of free Met, which competes with Lys for Asp and Glu as precursors, was increased unexpectedly by up to approximately 38-fold in the various transgenic and knockout plants. Together, our results show that Lys catabolism plays a major regulatory role in Lys accumulation in Arabidopsis seeds and reveal novel regulatory networks of seed amino acid metabolism. 相似文献
11.
12.
Mamputha S Lu ZL Roeske RW Millar RP Katz AA Flanagan CA 《Molecular endocrinology (Baltimore, Md.)》2007,21(1):281-292
GnRH I regulates reproduction. A second form, designated GnRH II, selectively binds type II GnRH receptors. Amino acids of the type I GnRH receptor required for binding of GnRH I (Asp2.61(98), Asn2.65(102), and Lys3.32(121)) are conserved in the type II GnRH receptor, but their roles in receptor function are unknown. We have delineated their functions using mutagenesis, signaling and binding assays, immunoblotting, and computational modeling. Mutating Asp2.61(97) to Glu or Ala, Asn2.65(101) to Ala, or Lys3.32(120) to Gln decreased potency of GnRH II-stimulated inositol phosphate production. Consistent with proposed roles in ligand recognition, mutations eliminated measurable binding of GnRH II, whereas expression of mutant receptors was not decreased. In detailed analysis of how these residues affect ligand-dependent signaling, [Trp2]-GnRH I showed lesser decreases in potency than GnRH I at the Asp2.61(97)Glu mutant. In contrast, [Trp2]-GnRH II showed the same loss of potency as GnRH II at this mutant. This suggests that Asp2.61(97) contributes to recognition of His2 of GnRH I, but not of GnRH II. GnRH II showed a large decrease in potency at the Asn2.65(101)Ala mutant compared with analogs lacking the CO group of Gly10NH2. This suggests that Asn2.65(101) recognizes Gly10NH2 of GnRH II. GnRH agonists showed large decreases in potency at the Lys3.32(120)Gln mutant, but antagonist activity was unaffected. This suggests that Lys3.32(120) recognizes agonists, but not antagonists, as in the type I receptor. These data indicate that roles of conserved residues are similar, but not identical, in the type I and II GnRH receptors. 相似文献
13.
Proteins from thermophiles are more stable than those from mesophiles. Several factors have been suggested as causes for this greater stability, but no general rule has been found. The amino acid composition of thermophile proteins indicates that the content of polar amino acids such as Asn, Gln, Ser, and Thr is lower, and that of charged amino acids such as Arg, Glu, and Lys is higher than in mesophile proteins. Among charged amino acids, however, the content of Asp is even lower in thermophile proteins than in mesophile proteins. To investigate the reasons for the lower occurrence of Asp compared to Glu in thermophile proteins, Glu was substituted with Asp in a hyperthermophile protein, MjTRX, and Asp was substituted with Glu in a mesophile protein, ETRX. Each substitution of Glu with Asp decreased the Tm of MjTRX by about 2 degrees C, while each substitution of Asp with Glu increased the Tm of ETRX by about 1.5 degrees C. The change of Tm destabilizes the MjTRX by 0.55 kcal/mol and stabilizes the ETRX by 0.45 kcal/mol in free energy. 相似文献
14.
Severity of experimental traumatic brain injury modulates changes in concentrations of cerebral free amino acids
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Valentina Di Pietro Stefano Signoretti Giuseppe Lazzarino Antonio Belli Barbara Tavazzi 《Journal of cellular and molecular medicine》2017,21(3):530-542
In this study, concentrations of free amino acids (FAA) and amino group containing compounds (AGCC) following graded diffuse traumatic brain injury (mild TBI, mTBI; severe TBI, sTBI) were evaluated. After 6, 12, 24, 48 and 120 hr aspartate (Asp), glutamate (Glu), asparagine (Asn), serine (Ser), glutamine (Gln), histidine (His), glycine (Gly), threonine (Thr), citrulline (Cit), arginine (Arg), alanine (Ala), taurine (Tau), γ‐aminobutyrate (GABA), tyrosine (Tyr), S‐adenosylhomocysteine (SAH), l ‐cystathionine (l ‐Cystat), valine (Val), methionine (Met), tryptophane (Trp), phenylalanine (Phe), isoleucine (Ile), leucine (Leu), ornithine (Orn), lysine (Lys), plus N‐acetylaspartate (NAA) were determined in whole brain extracts (n = 6 rats at each time for both TBI levels). Sham‐operated animals (n = 6) were used as controls. Results demonstrated that mTBI caused modest, transient changes in NAA, Asp, GABA, Gly, Arg. Following sTBI, animals showed profound, long‐lasting modifications of Glu, Gln, NAA, Asp, GABA, Ser, Gly, Ala, Arg, Citr, Tau, Met, SAH, l ‐Cystat, Tyr and Phe. Increase in Glu and Gln, depletion of NAA and Asp increase, suggested a link between NAA hydrolysis and excitotoxicity after sTBI. Additionally, sTBI rats showed net imbalances of the Glu‐Gln/GABA cycle between neurons and astrocytes, and of the methyl‐cycle (demonstrated by decrease in Met, and increase in SAH and l ‐Cystat), throughout the post‐injury period. Besides evidencing new potential targets for novel pharmacological treatments, these results suggest that the force acting on the brain tissue at the time of the impact is the main determinant of the reactions ignited and involving amino acid metabolism. 相似文献
15.
Masayochi Iwaki Hiroyuki Kagamiyama Mitsuhiro Nozaki 《Archives of biochemistry and biophysics》1981,210(1):210-223
The complete amino acid sequence of the β-subunit of protocatechuate 3,4-dioxygenase was determined. The β-subunit contained four methionine residues. Thus, five peptides were obtained after cleavage of the carboxymethylated β-subunit with cyanogen bromide, and were isolated on Sephadex G-75 column chromatography. The amino acid sequences of the cyanogen bromide peptides were established by characterization of the peptides obtained after digestion with trypsin, chymotrypsin, thermolysin, or Staphylococcus aureus protease. The major sequencing techniques used were automated and manual Edman degradations. The five cyanogen bromide peptides were aligned by means of the amino acid sequences of the peptides containing methionine purified from the tryptic hydrolysate of the carboxymethylated β-subunit. The amino acid sequence of all the 238 residues was as follows: ProAlaGlnAspAsnSerArgPheValIleArgAsp ArgAsnTrpHis ProLysAlaLeuThrPro-Asp — TyrLysThrSerIleAlaArg SerProArgGlnAla LeuValSerIleProGlnSer — IleSerGluThrThrGly ProAsnPheSerHisLeu GlyPheGlyAlaHisAsp-His — AspLeuLeuLeuAsnPheAsn AsnGlyGlyLeu ProIleGlyGluArgIle-Ile — ValAlaGlyArgValValAsp GlnTyrGlyLysPro ValProAsnThrLeuValGluMet — TrpGlnAlaAsnAla GlyGlyArgTyrArg HisLysAsnAspArgTyrLeuAlaPro — LeuAspProAsn PheGlyGlyValGly ArgCysLeuThrAspSerAspGlyTyrTyr — SerPheArg ThrIleLysProGlyPro TyrProTrpArgAsnGlyProAsnAsp — TrpArgProAla HisIleHisPheGlyIle SerGlyProSerIleAlaThr-Lys — LeuIleThrGlnLeuTyr PheGluGlyAspPro LeuIleProMetCysProIleVal — LysSerIleAlaAsn ProGluAlaValGlnGln LeuIleAlaLysLeuAspMetAsnAsn — AlaAsnProMet AsnCysLeuAlaTyr ArgPheAspIleValLeuArgGlyGlnArgLysThrHis PheGluAsnCys. The sequence published earlier in summary form (Iwaki et al., 1979, J. Biochem.86, 1159–1162) contained a few errors which are pointed out in this paper. 相似文献
16.
Amino acid sequence of a protease inhibitor isolated from Sarcophaga bullata determined by mass spectrometry. 总被引:2,自引:2,他引:0
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I. A. Papayannopoulos K. Biemann 《Protein science : a publication of the Protein Society》1992,1(2):278-288
The amino acid sequence of a protease inhibitor isolated from the hemolymph of Sarcophaga bullata larvae was determined by tandem mass spectrometry. Homology considerations with respect to other protease inhibitors with known primary structures assisted in the choice of the procedure followed in the sequence determination and in the alignment of the various peptides obtained from specific chemical cleavage at cysteines and enzyme digests of the S. bullata protease inhibitor. The resulting sequence of 57 residues is as follows: Val Asp Lys Ser Ala Cys Leu Gln Pro Lys Glu Val Gly Pro Cys Arg Lys Ser Asp Phe Val Phe Phe Tyr Asn Ala Asp Thr Lys Ala Cys Glu Glu Phe Leu Tyr Gly Gly Cys Arg Gly Asn Asp Asn Arg Phe Asn Thr Lys Glu Glu Cys Glu Lys Leu Cys Leu. 相似文献
17.
Arkadii V. Tarasevych Alexander E. Sorochinsky Valery P. Kukhar Jean-Claude Guillemin 《Origins of life and evolution of the biosphere》2013,43(2):129-135
Deracemization of a 50/50 mixture of enantiomers of aliphatic amino acids (Ala, Leu, Pro, Val) can be achieved by a simple sublimation of a pre-solubilized solid mixture of the racemates with a huge amount of a less-volatile optically active amino acid (Asn, Asp, Glu, Ser, Thr). The choice of chirality correlates with the handedness of the enantiopure amino acids—Asn, Asp, Glu, Ser, and Thr. The deracemization, enantioenrichment and enantiodepletion observed in these experiments clearly demonstrate the preferential homochiral interactions and a tendency of natural amino acids to homochiral self-organization. These data may contribute toward an ultimate understanding of the pathways by which prebiological homochirality might have emerged. 相似文献
18.
Nakashima K Ishida A Yakabe Y Yamazaki M Abe H 《Bioscience, biotechnology, and biochemistry》2006,70(8):1975-1978
We examined the effects of orally administrated amino acids on myfibrillar proteolysis in food-deprived chicks. Plasma N(tau)-methylhistidine concentration, as an index of myofibrillar proteolysis, was decreased by the administration of Glu, Gly, Ala, Leu, Ile, Ser, Thr, Met, Trp, Asn, Gln, Pro, Lys and Arg but not by Asp, Val, Phe, Tyr or His to chicks. Orally administrated Cys was fatal to chicks. These results indicate that oral Glu, Gly, Ala, Leu, Ile, Ser, Thr, Met, Trp, Asn, Gln, Pro, Lys and Arg administration suppressed myofibrillar proteolysis in chicks. 相似文献
19.
C. Q. Pan J. S. Ulmer A. Herzka R. A. Lazarus 《Protein science : a publication of the Protein Society》1998,7(3):628-636
Human deoxyribonuclease I (DNase I), an enzyme used to treat cystic fibrosis patients, has been systematically analyzed by site-directed mutagenesis of residues at the DNA binding interface. Crystal structures of bovine DNase I complexed with two different oligonucleotides have implicated the participation of over 20 amino acids in catalysis or DNA recognition. These residues have been classified into four groups based on the characterization of over 80 human DNase I variants. Mutations at any of the four catalytic amino acids His 134, His 252, Glu 78, and Asp 212 drastically reduced the hydrolytic activity of DNase I. Replacing the three putative divalent metal ion-coordinating residues Glu 39, Asp 168, or Asp 251 led to inactive variants. Amino acids Gln 9, Arg 41, Tyr 76, Arg 111, Asn 170, Tyr 175, and Tyr 211 were also critical for activity, presumably because of their close proximity to the active site, while more peripheral DNA interactions stemming from 13 other positions were of minimal significance. The relative importance of these 27 positions is consistent with evolutionary relationships among DNase I across different species, DNase I-like proteins, and bacterial sphingomyelinases, suggesting a fingerprint for a family of DNase I-like proteins. Furthermore, we found no evidence for a second active site that had been previously implicated in Mn2+-dependent DNA degradation. Finally, we correlated our mutational analysis of human DNase I to that of bovine DNase I with respect to their specific activity and dependence on divalent metal ions. 相似文献
20.
Amino Acids in Rat Striatal Dialysates: Methodological Aspects and Changes After Electroconvulsive Shock 总被引:11,自引:11,他引:0
Extracellular levels of amino acids were estimated in dialysates of the rat striatum that were collected 1, 2, and/or more than 5 days after surgery, before (resting release) and during exposure to high K concentrations (50 mM) or electroconvulsive shocks. The resting release of several amino acids (Glu, Asn, Thr, Tau, Tyr, Gly, and Ala) was higher 9 days as compared to 1 day after surgery. In the 1-day preparation the resting release correlated highly with that observed with push-pull cannulas. The correlation with the tissue content of the amino acids was high only when they were divided into two groups (putative transmitters and metabolic intermediates). High K exposure produced increased output of Ala, ethanolamine (Eam), Asp, Glu, Tau, and Gly and a decrease in the egress of Gln 1 or 2 days after surgery. The effects on Asp and Glu had disappeared, and that on Gln reversed after 4-9 days. Electrically induced convulsions produced increased output of Ala, Gln, and Eam 1 or 2 days and 2 weeks after implantation of the probe. Changes were seen not only during but also (and some cases even more prominent) after the seizure. This study shows the usefulness of dialysis to monitor extracellular transmitter amino acids in the striatum of conscious rats (also bilateral dialysis was possible) for only a limited time after implantation of the probe. The dialysis method is suitable for longer time, when metabolic changes in amino acids are to be followed. In addition to transmitter release, glycolysis can be monitored by the measurement of Ala in the dialysate. 相似文献