共查询到20条相似文献,搜索用时 109 毫秒
1.
The response regulator DosR is essential for promoting long-term survival of Mycobacterium tuberculosis under low oxygen conditions in a dormant state and may be responsible for latent tuberculosis in one-third of the world's population. Here, we report crystal structures of full-length unphosphorylated DosR at 2.2 Å resolution and its C-terminal DNA-binding domain at 1.7 Å resolution. The full-length DosR structure reveals several features never seen before in other response regulators. The N-terminal domain of the full-length DosR structure has an unexpected (βα)4 topology instead of the canonical (βα)5 fold observed in other response regulators. The linker region adopts a unique conformation that contains two helices forming a four-helix bundle with two helices from another subunit, resulting in dimer formation. The C-terminal domain in the full-length DosR structure displays a novel location of helix α10, which allows Gln199 to interact with the catalytic Asp54 residue of the N-terminal domain. In contrast, the structure of the DosR C-terminal domain alone displays a remarkable unstructured conformation for helix α10 residues, different from the well-defined helical conformations in all other known structures, indicating considerable flexibility within the C-terminal domain. Our structures suggest a mode of DosR activation by phosphorylation via a helix rearrangement mechanism. 相似文献
2.
Hiroki Yamaguchi Akiko Kamegawa Kunio Nakata Tatsuki Kashiwagi Toshimi Mizukoshi Yoshinori Fujiyoshi Kazutoshi Tani 《Journal of structural biology》2019,205(1):11-21
Leucine dehydrogenase (LDH, EC 1.4.1.9) is a NAD+-dependent oxidoreductase that catalyzes the deamination of branched-chain l-amino acids (BCAAs). LDH of Geobacillus stearothermophilus (GstLDH) is a highly thermostable enzyme that has been applied for the quantification or production of BCAAs. Here the cryo-electron microscopy (cryo-EM) structures of apo and NAD+-bound LDH are reported at 3.0 and 3.2?Å resolution, respectively. On comparing the structures, the two overall structures are almost identical, but it was observed that the partial conformational change was triggered by the interaction between Ser147 and the nicotinamide moiety of NAD+. NAD+ binding also enhanced the strength of oligomerization interfaces formed by the core domains. Such additional interdomain interaction is in good agreement with our experimental results showing that the residual activity of NAD+-bound form was approximately three times higher than that of the apo form after incubation at 80?°C. In addition, sequence comparison of three structurally known LDHs indicated a set of candidates for site-directed mutagenesis to improve thermostability. Subsequent mutation analysis actually revealed that non-conserved residues, including Ala94, Tyr127, and the C-terminal region, are crucial for oligomeric thermostability. 相似文献
3.
Arrangement of nucleotide sequences in adeno-associated virus DNA 总被引:22,自引:0,他引:22
There are two types of adeno-associated virus virions which contain complementary single-stranded DNA genomes of about 1.4 × 106 daltons. The purified complementary single polynucleotide chains anneal to form duplex linear monomers, circular monomers, and linear dimers, in addition to other less well-defined structures, as identified by sedimentation and electron microscopy. All duplex species are formed by linear single polynucleotide chains of unit length, thus duplex circles and linear dimers are assumed to be held together by relatively short overlapping hydrogen-bonded regions. The initial linear monomers present after annealing the complementary single strands do not form duplex circles or oligomers when re-exposed to annealing conditions, but DNA which sediments as linear monomers after heating linear dimers at a temperature from 7 to 25 deg. C below the Tm of duplex AAV2 DNA does re-form oligomers and circles when exposed to annealing conditions. Denaturation and reannealing of any duplex species leads to the formation of all forms, indicating that the over-all single strand composition of all species is equivalent. Disruption of duplex circles by limited exonucleolytic digestion using either 3′ or 5′ exonucleases leads to the conclusion that the overlap region may have either 3′ or 5′ termini and that the overlap region represents less than 6% of the length of the genome. Exonucleolytic digestion of linear monomers to the extent of 50% leaves polynucleotide chains which cannot reanneal after denaturation, thus AAV DNA is not randomly circularly permuted. Duplex linear monomers which do not form circles when exposed to annealing conditions do form duplex circles after 1% exonuclease III digestion. A model consistent with these data is one in which the linear single polynucleotide chains present in AAV virions consist of two or more permutations. All of these chains contain terminal repetitions and their start points occur within a limited region, representing < 6% of the length of the genome. According to this interpretation AAV DNA would exist within the virion as a linear single polynucleotide chain or is cleaved at a few specific sites during extraction. 相似文献
4.
Aaron P. Yamniuk Kathryn L. Anderson Marie E. Fraser Hans J. Vogel 《Protein science : a publication of the Protein Society》2009,18(5):1128-1134
Recent X-ray crystal structures and solution NMR spectroscopy data for calcium- and integrin-binding protein 1 (CIB1) have all revealed a common EF-hand domain structure for the protein. However, the orientation of the two protein domains, the oligomerization state, and the conformations of the N- and C-terminal extensions differ among the structures. In this study, we examine whether the binding of glutathione or auxiliary Ca2+ ions as observed in the crystal structures, occur in solution, and whether these interactions can influence the structure or dimerization of CIB1. In addition, we test the potential phosphatase activity of CIB1, which was hypothesized based on the glutathione binding site geometry observed in one of the crystal structures of the protein. Biophysical and biochemical experiments failed to detect glutathione binding, protein dimerization, or phosphatase activity for CIB1 under several solution conditions. However, our data identify low affinity (Kd, 10−2M) Ca2+ binding events that influence the structures of the N- and C-terminal extensions of CIB1 under high (300 mM) Ca2+ crystallization conditions. In addition to providing a rationale for differences amongst the various solution and crystal structures of CIB1, our results show that the impact of low affinity Ca2+ binding events should be considered when analyzing and interpreting protein crystallographic structures determined in the presence of very high Ca2+ concentrations. 相似文献
5.
I. P. Yermakova T. V. Potanina V. P. Busulina I. A. Pronchenko I. M. Larina N. P. Shmerko V. I. Sevastianov 《Human physiology》2013,39(2):184-189
The kinetics of calcium binding by tissue structures was analyzed in 19 healthy volunteers (13 men and 6 women) from the age group of 33 ± 6.5 years under the conditions of acute hypercalcemia followed by drip intravenous infusion of calcium gluconate for 2.5 h. At the end of each 30-min period, the calcium amount retained by the tissue structures was recorded and the kinetic parameters of calcium binding were determined according to Langmuir and Scatchard. In all volunteers, there was a region of binding isotherm with positive cooperativity (linear regression in Scatchard’s coordinates) with the same buffer capacity (βtis) for calcium in Langmuir’s coordinates (0.58 ± 0.24 L kg). Half of the volunteers exhibited cooperativity at [Ca2+] = 1.3–1.5 mmol/L, while others, at [Ca2+] = 1.0–1.3 mmol/L, which corresponded to the differences in the association constant (Ka) and the number of interactive sites (n) with [Ca2+] = 1 mmol/L. Additionally, two segments of the binding isotherm were detected with the successive binding of calcium to one set of non-interactive centers with similar kinetic parameters of calcium binding (βtis, Ka, n). Four different types of calcium binding in healthy volunteers were found. The results of this study may serve as the basis for a functional diagnostic test of disorders of the tissue calcium-binding properties in different pathological conditions. 相似文献
6.
Joon Ki Hong Jihyun Je Chieun Song Jung Eun Hwang Yeon-Hee Lee Chae Oh Lim 《Genes & genomics.》2012,34(1):13-18
The phytocystatins are inhibitors of papain-like cysteine proteinases that are implicated in defense mechanisms and the regulation of protein turnover. BCPI-1, a Brassica rapa (Chinese cabbage) phytocystatin isolated from flower buds, contains an extended C-terminal region that contains a single Cys residue at position 102. In an effort to investigate the role of the C-terminus and this Cys residue in BCPI-1 activity, purified recombinant proteins of BCPI-1, including wild-type BCPI-1 (wtBCPI-1), N-terminus BCPI-1 (BCPI-1??C), C-terminus BCPI-1 (BCPI-1??N), and BCPI-1 with a single Cys residue exchange to Ser (BCPI-1C102S), were generated and their inhibitory activities against papain were investigated. Kinetic analysis revealed that the monomeric forms of wtBCPI-1 (K i = 6.84 ± 0.3 × 10?8 M) inhibited papain more efficiently than the dimeric forms of wtBCPI-1 (K i = 1.01 ± 0.5 × 10?7 M). Experiments with recombinant BCPI-1C102S demonstrated that the dimerization of wtBCPI-1 caused by the formation of an intermolecular disulfide bond at the cysteine residue. The inhibitory activity of the recombinant proteins, except BCPI-1??N, was reduced in the pH range of 7.0?C11.5 and was highly stable over a wide range of temperatures. Thus, dimerization mediated by the cysteine residue in the extended C-terminal region and alkaline conditions reduced the inhibitory activity of BCPI-1. 相似文献
7.
《MABS-AUSTIN》2013,5(7):1289-1299
ABSTRACTIgG4, a common type of therapeutic antibody, is less stable during manufacturing processes compared with IgG1. Aggregation and fragmentation are the two main challenges. Here, we report instability of the heavy chain (HC) C-terminal region under acidic conditions, which leads to cleavage and aggregation. Leu445, at the C-terminal region of the HC in IgG4, plays a critical role in its acid-induced fragmentation and subsequent aggregation. We found that mutating HC C-terminal Leu445 to Pro (the corresponding residue in IgG1) in IgG4_CDR-X significantly reduces fragmentation and aggregation, while mutating Pro445 to Leu in IgG1_CDR-X promotes fragmentation and aggregation. HC C-terminal Gly446 cleavage was observed in low pH citrate buffer and resulted in further fragmentation and aggregation, whereas, glycine buffer can completely inhibit the cleavage and aggregation. It is proposed that cleavages occur through acid-induced hydrolysis under acidic conditions and glycine stabilizes IgG4 via two main mechanisms: 1) product feedback inhibition of the hydrolysis reaction, and 2) stabilization of protein conformation by direct interaction with the peptide backbone and charged side chains. Experiments using IgG4 molecules IgG4_CDR-Y and IgG4_CDR-Z with the same CH domains as IgG4_CDR-X, but different complementarity-determining regions (CDRs), indicate that the stability of the HC C-terminal region is also closely related to the sequence of the CDRs. The stability of IgG4_CDR-X is significantly improved when binding to its target. Both observations suggest that there are potential interactions between Fab and CH2-CH3 domains, which could be the key factor affecting the stability of IgG antibodies. 相似文献
8.
Biljan I Ilc G Giachin G Raspadori A Zhukov I Plavec J Legname G 《Journal of molecular biology》2011,412(4):660-673
The development of transmissible spongiform encephalopathies (TSEs) is associated with the conversion of the cellular prion protein (PrPC) into a misfolded, pathogenic isoform (PrPSc). Spontaneous generation of PrPSc in inherited forms of disease is caused by mutations in gene coding for PrP (PRNP). In this work, we describe the NMR solution-state structure of the truncated recombinant human PrP (HuPrP) carrying the pathological V210I mutation linked to genetic Creutzfeldt-Jakob disease. The three-dimensional structure of V210I mutant consists of an unstructured N-terminal part (residues 90-124) and a well-defined C-terminal domain (residues 125-228). The C-terminal domain contains three α-helices (residues 144-156, 170-194 and 200-228) and a short antiparallel β-sheet (residues 129-130 and 162-163). Comparison with the structure of the wild-type HuPrP revealed that although two structures share similar global architecture, mutation introduces some local structural differences. The observed variations are mostly clustered in the α2-α3 inter-helical interface and in the β2-α2 loop region. Introduction of bulkier Ile at position 210 induces reorientations of several residues that are part of hydrophobic core, thus influencing α2-α3 inter-helical interactions. Another important structural feature involves the alteration of conformation of the β2-α2 loop region and the subsequent exposure of hydrophobic cluster to solvent, which facilitates intermolecular interactions involved in spontaneous generation of PrPSc. The NMR structure of V210I mutant offers new clues about the earliest events of the pathogenic conversion process that could be used for the development of antiprion drugs. 相似文献
9.
A. Higashitani Y. Ishii Y. Kato K. Horiuchi 《Molecular genetics and genomics : MGG》1997,254(4):351-357
SulA is induced in Escherichia coli by the SOS response and inhibits cell division through interaction with FtsZ. To determine which region of SulA is essential for the inhibition of cell division, we constructed a series of N-terminal and C-terminal deletions of SulA and a series of alanine substitution mutants. Arginine at position 62, leucine at 67, tryptophan at 77 and lysine at 87, in the central region of SulA, were all essential for the inhibitory activity. Residues 3–27 and the C-terminal 21 residues were dispensable for the activity. The mutant protein lacking N-terminal residues 3–47 was inactive, as was that lacking the C-terminal 34 residues. C-terminal deletions of 8 and 21 residues increased the growth-inhibiting activity in lon + cells, but not in lon ? cells. The wild-type and mutant SulA proteins were isolated in a form fused to E. coli maltose-binding protein, and tested in vitro for sensitivity to Lon protease. Lon degraded wild-type SulA and a deletion mutant lacking the N-terminal 93 amino acids, but did not degrade the derivative lacking 21 residues at the C-terminus. Futhermore, the wild-type SulA and the N-terminal deletion mutant formed a stable complex with Lon, while the C-terminal deletion did not. MBP fused to the C-terminal 20 residues of SulA formed a stable complex with, but was not degraded by Lon. When LacZ protein was fused at its C-terminus to 8 or 20 amino acid residues from the C-terminal region of SulA the protein was stable in lon + cells. These results indicate that the C-terminal 20 residues of SulA permit recognition by, and complex formation with, Lon, and are necessary, but not sufficient, for degradation by Lon. 相似文献
10.
Morozumi N Hanada T Habara H Yamaki A Furuya M Nakatsuka T Inomata N Minamitake Y Ohsuye K Kangawa K 《Peptides》2011,32(5):1001-1007
Ghrelin is an endogenous ligand for growth hormone secretagogue receptor 1a (GHS-R1a), and consists of 28 amino acid residues with octanoyl modification at Ser3. The previous studies have revealed that N-terminal part of ghrelin including modified Ser3 is the active core for the activation of GHS-R1a. On the other hand, the role of C-terminal (8-28) region in ghrelin has not been clarified yet. In the present study, we prepared human ghrelin, C-terminal truncated ghrelin derivatives and anamorelin, a small molecular GHS compound which supposedly mimics the N-terminal active core, and examined GHS-R1a agonist activity in vitro, pharmacokinetic (PK) profile and growth hormone (GH) releasing activity in rats. All compounds demonstrated potent GHS-R1a agonist activities in vitro. Although the lack of C-terminal two amino acids did not modify PK profile and GH releasing activity, the deletion of C-terminal 8 and 20 amino acids affected them, and ghrelin(1-7)-Lys-NH2 exhibited very short plasma half-life and low GH releasing activity in vivo. In rat plasma, ghrelin(1-7)-Lys-NH2 was degraded more rapidly than ghrelin, suggesting that C-terminal part of ghrelin protected octanoylation of Ser3 from plasma esterases. Subdiaphragmatic vagotomy significantly attenuated GH response to ghrelin but not to anamorelin. These results suggest that the C-terminal part of ghrelin has an important role in the biological activity in vivo. We also found that ghrelin stimulated GH release mainly via a vagal nerve pathway but anamorelin augmented GH release possibly by directly acting on brain in rats. 相似文献
11.
The transport of calcium to the extracellular space carried out by plasma membrane Ca2+ pumps (PMCAs) is essential for maintaining low Ca2+ concentrations in the cytosol of eukaryotic cells. The activity of PMCAs is controlled by autoinhibition. Autoinhibition is relieved by the binding of Ca2+-calmodulin to the calmodulin-binding autoinhibitory sequence, which in the human PMCA is located in the C-terminal segment and results in a PMCA of high maximal velocity of transport and high affinity for Ca2+. Autoinhibition involves the intramolecular interaction between the autoinhibitory domain and a not well defined region of the molecule near the catalytic site. Here we show that the fusion of GFP to the C terminus of the h4xb PMCA causes partial loss of autoinhibition by specifically increasing the Vmax. Mutation of residue Glu99 to Lys in the cytosolic portion of the M1 transmembrane helix at the other end of the molecule brought the Vmax of the h4xb PMCA to near that of the calmodulin-activated enzyme without increasing the apparent affinity for Ca2+. Altogether, the results suggest that the autoinhibitory interaction of the extreme C-terminal segment of the h4 PMCA is disturbed by changes of negatively charged residues of the N-terminal region. This would be consistent with a recently proposed model of an autoinhibited form of the plant ACA8 pump, although some differences are noted. 相似文献
12.
Wolfgang Haehnel 《BBA》1982,682(2):245-257
Signal I, the EPR signal of P-700, induced by long flashes as well as the rate of linear electron transport are investigated at partial inhibition of electron transport in chloroplasts. Inhibition of plastoquinol oxidation by dibromothymoquinone and bathophenanthroline, inhibition of plastocyanin by KCN and HgCl2, and inhibition by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide are used to study a possible electron exchange between electron-transport chains after plastoquinone. (1) At partial inhibition of plastocyanin the reduction kinetics of P-700+ show a fast component comparable to that in control chloroplasts and a new slow component. The slow component indicates P-700+ which is not accessible to residual active plastocyanin under these conditions. We conclude that P-700 is reduced via complexed plastocyanin. (2) The rate of linear electron transport at continuous illumination decreases immediately when increasing amounts of plastocyanin are inhibited by KCN incubation. This is not consistent with an oxidation of cytochrome f by a mobile pool of plastocyanin with respect to the reaction rates of plastocyanin being more than an order of magnitude faster than the rate-limiting step of linear electron transport. It is evidence for a complex between the cytochrome b6 - f complex and plastocyanin. The number of these complexes with active plastocyanin is concluded to control the rate-limiting plastoquinol oxidation. (3) Partial inhibition of the electron transfer between plastoquinone and cytochrome f by dibromothymoquinone and bathophenanthroline causes decelerated monophasic reduction of total P-700+. The P-700 kinetics indicate an electron transfer from the cytochrome b6 - f complex to more than ten Photosystem I reaction center complexes. This cooperation is concluded to occur by lateral diffusion of both complexes in the membrane. (4) The proposed functional organization of electron transport from plastoquinone to P-700 in situ is supported by further kinetic details and is discussed in terms of the spatial distribution of the electron carriers in the thylakoid membrane. 相似文献
13.
Mouilleron S Badet-Denisot MA Golinelli-Pimpaneau B 《Journal of molecular biology》2008,377(4):1174-1185
Glucosamine-6-phosphate synthase (GlmS) channels ammonia from glutamine at the glutaminase site to fructose 6-phosphate (Fru6P) at the synthase site. Escherichia coli GlmS is composed of two C-terminal synthase domains that form the dimer interface and two N-terminal glutaminase domains at its periphery. We report the crystal structures of GlmS alone and in complex with the glucosamine-6-phosphate product at 2.95 Å and 2.9 Å resolution, respectively. Surprisingly, although the whole protein is present in this crystal form, no electron density for the glutaminase domain was observed, indicating its mobility. Comparison of the two structures with that of the previously reported GlmS-Fru6P complex shows that, upon sugar binding, the C-terminal loop, which forms the major part of the channel walls, becomes ordered and covers the synthase site. The ordering of the glutaminase domains likely follows Fru6P binding by the anchoring of Trp74, which acts as the gate of the channel, on the closed C-terminal loop. This is accompanied by a major conformational change of the side chain of Lys503# of the neighboring synthase domain that strengthens the interactions of the synthase domain with the C-terminal loop and completely shields the synthase site. The concomitant conformational change of the Lys503#-Gly505# tripeptide places catalytic His504# in the proper position to open the sugar and buries the linear sugar, which is now in the vicinity of the catalytic groups involved in the sugar isomerization reaction. Together with the previously reported structures of GlmS in complex with Fru6P or glucose 6-phosphate and a glutamine analogue, the new structures reveal the structural changes occurring during the whole catalytic cycle. 相似文献
14.
Interaction between the C-terminal domains of N and P proteins of measles virus investigated by NMR 总被引:1,自引:0,他引:1
In this paper we investigate the interaction between the C-terminal domains of the measles virus phosphoprotein (XD) and nucleoprotein (NTAIL) by using nuclear magnetic resonance chemical shift perturbation experiments. Using both NTAIL constructs and peptides, we show that contrary to the conserved Box2 region (N489-506), the C-terminal region of NTAIL (N513-525) does not directly interact with XD, and yet affects binding to XD. We tentatively propose a model where the C-terminus of NTAIL would stabilize the NTAIL-XD complex either via a functional coupling with N489-506 or by reducing the entropic penalty associated to the binding-coupled-to-folding process.
Structured summary
MINT-7009780, MINT-7009793, MINT-7009808: N-tail (uniprotkb:Q89933) and P (uniprotkb:P03422) bind (MI:0407) by nuclear magnetic resonance (MI:0077) 相似文献15.
Structural and biochemical analyses of hemimethylated DNA binding by the SeqA protein 总被引:1,自引:1,他引:0
Fujikawa N Kurumizaka H Nureki O Tanaka Y Yamazoe M Hiraga S Yokoyama S 《Nucleic acids research》2004,32(1):82-92
The Escherichia coli SeqA protein recognizes the 11 hemimethylated G-mA-T-C sites in the oriC region of the chromosome, and prevents replication over-initiation within one cell cycle. The crystal structure of the SeqA C-terminal domain with hemimethylated DNA revealed the N6-methyladenine recognition mechanism; however, the mechanism of discrimination between the hemimethylated and fully methylated states has remained elusive. In the present study, we performed mutational analyses of hemimethylated G-mA-T-C sequences with the minimal DNA-binding domain of SeqA (SeqA71–181), and found that SeqA71–181 specifically binds to hemimethylated DNA containing a sequence with a mismatched mA:G base pair [G-mA(:G)-T-C] as efficiently as the normal hemimethylated G-mA(:T)-T-C sequence. We determined the crystal structures of SeqA71–181 complexed with the mismatched and normal hemimethylated DNAs at 2.5 and 3.0 Å resolutions, respectively, and found that the mismatched mA:G base pair and the normal mA:T base pair are recognized by SeqA in a similar manner. Furthermore, in both crystal structures, an electron density is present near the unmethylated adenine, which is only methylated in the fully methylated state. This electron density, which may be due to a water molecule or a metal ion, can exist in the hemimethylated state, but not in the fully methylated state, because of steric clash with the additional methyl group. 相似文献
16.
Johnny Stiban Gregory A. Farnum Stacy L. Hovde Laurie S. Kaguni 《The Journal of biological chemistry》2014,289(35):24032-24042
The metazoan mitochondrial DNA helicase is an integral part of the minimal mitochondrial replisome. It exhibits strong sequence homology with the bacteriophage T7 gene 4 protein primase-helicase (T7 gp4). Both proteins contain distinct N- and C-terminal domains separated by a flexible linker. The C-terminal domain catalyzes its characteristic DNA-dependent NTPase activity, and can unwind duplex DNA substrates independently of the N-terminal domain. Whereas the N-terminal domain in T7 gp4 contains a DNA primase activity, this function is lost in metazoan mtDNA helicase. Thus, although the functions of the C-terminal domain and the linker are partially understood, the role of the N-terminal region in the metazoan replicative mtDNA helicase remains elusive. Here, we show that the N-terminal domain of Drosophila melanogaster mtDNA helicase coordinates iron in a 2Fe-2S cluster that enhances protein stability in vitro. The N-terminal domain binds the cluster through conserved cysteine residues (Cys68, Cys71, Cys102, and Cys105) that are responsible for coordinating zinc in T7 gp4. Moreover, we show that the N-terminal domain binds both single- and double-stranded DNA oligomers, with an apparent Kd of ∼120 nm. These findings suggest a possible role for the N-terminal domain of metazoan mtDNA helicase in recruiting and binding DNA at the replication fork. 相似文献
17.
The gene (1,542 bp) encoding thermostable Ca2+-independent and raw starch hydrolyzing α-amylase of the extremely thermophilic bacterium Geobacillus thermoleovorans encodes for a protein of 50 kDa (Gt-amyII) with 488 amino acids. The enzyme is optimally active at pH 7.0 and 60 °C with a t 1/2 of 19.4 h at 60 and 4 h at 70 °C. Gt-amyII hydrolyses corn and tapioca raw starches efficiently and therefore finds application in starch saccharification at industrial sub-gelatinisation temperatures. The starch hydrolysis is facilitated following adsorption of the enzyme to starch at the C-terminal domain, as confirmed by the truncation analysis. The adsorption rate constant of Gt-amyII to raw corn starch is 37.6-fold greater than that for the C-terminus truncated enzyme (Gt-amyII-T). Langmuir–Hinshelwood kinetic analysis in terms of equilibrium parameter (K R) suggested that the adsorption of Gt-amyII to corn starch is more favourable than that of Gt-amyII-T. Thermodynamics of temperature inactivation indicated a decrease in thermostabilisation of Gt-amyII upon truncation of its C-terminus. The addition of raw corn starch increased t 1/2 of Gt-amyII, but it has no such effect on Gt-amyII-T. It can, therefore, be stated that Gt-amyII binds to raw corn starch via C-terminal region that contributes to its thermostability. Phylogenetic analysis confirmed that starch binding region of Gt-amyII is, in fact, the non-catalytic domain C, and not the typical SBD of CBM families. The role of domain C in raw starch binding throws light on the evolutionary path of the known SBDs. 相似文献
18.
《Animal : an international journal of animal bioscience》2021,15(3):100142
Growing concerns regarding sustainability in agriculture include the availability of drinking water, which is putting pressure on livestock production, especially the beef sector, for more efficient practices. Thus, genetic parameters were estimated for traits related to water intake and water use efficiency in Senepol cattle. Senepol females (n = 925) and males (n = 191) were evaluated in performance tests carried out from 2014 to 2019. Daily dry matter intake (DMI) and water intake (WI) were recorded by electronic feed and water bunks (Intergado Ltd.). Other traits assessed included average daily gain (ADG); mid-test metabolic BW (BW0.75); residual water intake based on ADG (RWIADG), estimated as the residual of the linear regression equation of WI on ADG and BW0.75; residual water intake based on DMI (RWIDMI), estimated as the residual of the linear regression equation of WI on DMI and BW0.75 (RWIDMI); water conversion ratio (= WI/ADG); gross water efficiency (GWE = ADG/WI); residual feed intake estimated as the residual of the linear regression equation of DMI on ADG and BW0.75 (RFI); feed conversion ratio (= DMI/ADG) and gross feed efficiency. Genetic (co)variances were estimated with bivariate analyses. The heritabilities for WI, RWIADG and RWIDMI were 0.38, 0.36 and 0.33, respectively. Water conversion ratio, RWIADG and RWIDMI showed positive genetic and phenotypic correlations with WI, whereas GWE was negatively correlated with WI, suggesting that traits related to water use efficiency may be useful to identify cattle with reduced WI. Water intake showed positive genetic (r = 0.79) and phenotypic (r = 0.60) correlations with DMI, suggesting the use of WI to estimate DMI in future studies. Both RWIADG and RWIDMI were genetically correlated with RFI (0.67 and 0.57, respectively) and ADG (0.49 and 0.44, respectively), showing that RWI is positively associated with feed efficiency, but has an antagonistic relationship with growth. This antagonism, however, may be managed using selection indexes. Genetic improvement of water use efficiency in Senepol cattle is possible through selection and may reduce the water requirements of beef production systems. 相似文献
19.
N. V. Pletneva S. V. Pletnev A. M. Bogdanov E. A. Goryacheva I. V. Artemyev E. A. Suslova S. F. Arkhipova V. Z. Pletnev 《Russian Journal of Bioorganic Chemistry》2014,40(4):383-389
The spatial structure of dimeric green fluorescent protein EGFP-K162Q with MDELYK (EGFPv) C-terminal deletion has been assigned in the P61 space group with resolution 1.34 Å by X-ray diffraction analysis. The results have been compared with X-ray diffraction data of monomeric EGFP (green biomarker with enhanced photophysical properties) assigned in another crystal space group, P212121, with resolution 1.50 and 1.35 Å. Subunits in the EGFPv dimeric structure are located at 75° angle with the contact area ~800 Å2. The dimeric framework is stabilized by the six hydrogen bonds and central hydrophobic core of six residues. The root-mean-square deviation value for Cα atoms in 3–230 residues of the P61 and P212121 crystal structures is 0.55 Å. The differential characteristics of EGFPv-P61 structure, compared to that of P212121, is a noticeably different orientation of the Glu222 side chain, and a new conformation of the 155–159 loop fragment, characterized by deviations among the Cα atoms of superimposed structures reaching 4.6 Å for Lys156 and 5.5 Å for Lys158. 相似文献
20.
Prasad Kasaragod Rajaram Venkatesan Tiila R. Kiema J. Kalervo Hiltunen Rik K. Wierenga 《The Journal of biological chemistry》2010,285(31):24089-24098
The crystal structure of the full-length rat peroxisomal multifunctional enzyme, type 1 (rpMFE1), has been determined at 2.8 Å resolution. This enzyme has three catalytic activities and two active sites. The N-terminal part has the crotonase fold, which builds the active site for the Δ3,Δ2-enoyl-CoA isomerase and the Δ2-enoyl-CoA hydratase-1 catalytic activities, and the C-terminal part has the (3S)-hydroxyacyl-CoA dehydrogenase fold and makes the (3S)-hydroxyacyl-CoA dehydrogenase active site. rpMFE1 is a multidomain protein having five domains (A–E). The crystal structure of full-length rpMFE1 shows a flexible arrangement of the A-domain with respect to the B–E-domains. Because of a hinge region near the end of the A-domain, two different positions of the A-domain were observed for the two protein molecules (A and B) of the asymmetric unit. In the most closed conformation, the mode of binding of CoA is stabilized by domains A and B (helix-10), as seen in other crotonase fold members. Domain B, although functionally belonging to the N-terminal part, is found tightly associated with the C-terminal part, i.e. fixed to the E-domain. The two active sites of rpMFE1 are ∼40 Å apart, separated by a tunnel, characterized by an excess of positively charged side chains. Comparison of the structures of rpMFE1 with the monofunctional crotonase and (3S)-hydroxyacyl-CoA dehydrogenase superfamily enzymes, as well as with the bacterial α2β2-fatty acid oxidation multienzyme complex, reveals that this tunnel could be important for substrate channeling, as observed earlier on the basis of the kinetics of rpMFE1 purified from rat liver. 相似文献