Influence of selenium induced oxidative stress on spermatogenesis and lactate dehydrogenase-X in mice testis

Aim: To evaluate the effect of oxidative stress on the spermatogenesis and lactate dehydrogenase-X (LDH-X) activity in mouse testis. Methods: For creating different levels of oxidative stress in mice, three selenium (Se) level diets were fed in separate groups for 8 weeks. Group 1 animals were fed yeast-based Se-deficient (0.02 ppm) diet. Group 2 and Group 3 animals were fed with the same diet supplemented with 0.2 ppm and 1 ppm Se as sodium selenite, respectively. After 8 weeks, biochemical and histopathological observations of the testis were carried out. LDH-X levels in the testis were analyzed by western immunoblot and ELISA. Results: A significant decrease in testis Se level was observed in Group 1 animals, whereas it was enhanced in Group 3 as compared to Group 2. The glutathione peroxidase (GSH-Px) activity was significantly reduced in both the liver and testis in Group 1, but not in Group 2 and 3. A significant increase in the testis glutathione-S-transferase (GST) activity was observed in Group 1, whereas no significant change was seen in Groups 2 and 3. Histological analysis of testis revealed a normal structure in Group 2. A significant decrease in the germ cell population in Group 1 was observed as compared to Group 2 with the spermatids and mature sperm affected the most. Decrease in the lumen size was also observed. In the Se-excess group (Group 3), displacement of germ cell population was observed. Further, a decrease in the LDH-X level in testis was observed in Group 1. Conclusion: Excessive oxidative stress in the Se deficient group, as indicated by changes in the GSH-Px/GST activity, affects the spermatogenic process with a reduction in mature sperm and in turn the LDH-X level.

Systemic immune inflammation index, ratio of lymphocytes to monocytes, lactate dehydrogenase and prognosis of diffuse large Bcell lymphoma patients

BACKGROUND In malignant tumors,inflammation plays a vital role in the development,invasion,and metastasis of cancer cells.Diffuse large B-cell lymphoma(DLBCL),the most common malignant proliferative disease of the lymphatic system,is commonly associated with inflammation.The international prognostic index(IPI),which includes age,lactate dehydrogenase(LDH),number of extranodal lesions,Ann Arbor score,and Eastern Cooperative Oncology Group(ECOG)score,can evaluate the prognosis of DLBCL.However,its use in accurately identifying highrisk patients and guiding treatment is poor.Therefore,it is important to find novel immune markers in predicting the prognosis of DLBCL patients.AIM To determine the association between the systemic immune inflammation index(SII),ratio of lymphocytes to monocytes(LMR),ratio of LMR to LDH(LMR/LDH),and prognosis of patients with DLBCL.METHODS A total of 68 patients diagnosed with DLBCL,treated in our hospital between January 2016 and January 2020,were included.χ2 test,Pearson’s R correlation,Kaplan Meier curves,and Cox proportional risk regression analysis were used.The differences in the SII,LMR,and LMR/LDH among patients with different clinicopathological features were analyzed.The differences in progression-free survival time among patients with different SII,LMR,and LMR/LDH expressions and influencing factors affecting the prognosis of DLBCL patients,were also analyzed.RESULTS The LMR and LMR/LDH in patients with Ann Arbor stage III–IV,ECOG score≥2,and SII,IPI score 2–5 were significantly higher than those of patients with Ann Arbor stage I-II and ECOG score<2(P<0.05).Patients with high SII,LMR,and LMR/LDH had progression-free survival times of 34 mo(95%CI:32.52–38.50),35 mo(95%CI:33.42–36.58)and 35 mo(95%CI:33.49–36.51),respectively,which were significantly lower than those with low SII,LMR,and LMR/LDH(P<0.05);the SII,LMR,and LMR/LDH were positively correlated(P<0.05).Cox proportional risk regression analysis showed that the SII,LMR,and LMR/LDH were influencing factors for the prognosis of DLBCL patients(hazard ratio=1.143,1.665,and 1.704,respectively;P<0.05).CONCLUSION The SII,LMR,and LMR/LDH are related to the clinicopathological features of DLCBL,and they also influence the prognosis of patients with the disease.

Malate and Lactate Dehydrogenases of a Freshwater Catfish: Impact of Endosulfan

A sublethal concentration of technical grade endosulfan (END) inhibited 35 to 55% of the activities of cytoplasmic malate dehydrogenase (cMDH), mitochondrial malate dehydrogenase (mMDH), and lactate dehydrogenase (LDH) in the liver and the skeletal muscle of a freshwater catfish, Clarias halrachus, after 7 days of exposure. The activity remained in the inhibited state up to 28 days. The withdrawal of END from the medium after 1 week of exposure gradually restored the activities to control levels within 21 days in the skeletal muscle and 28 days in the liver. The administration of actinomycin D or cycloheximide between the 14th and the 21st day of the withdrawal of END almost completely inhibited the withdrawal-dependent recovery in the activities of all the three enzymes. This indicates de novo synthesis of the enzymes during the recovery period. A conjoint treatment of END and triiodothyronine (T_3) raised the activities of cMDH, mMDH, and LDH in the liver and the skeletal muscle up to the control levels. This shows that the inhibitory effect of END may be relieved in presence of T_3. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed few changes in the pattern of cytoplasmic proteins of the liver and the skeletal muscle in response to exposure to END. 1990 Academic Press. Inc.

Function of Lactate Dehydrogenase in Cardiac and Skeletal Muscle of Phrynocephalus Lizard in Relation to High-Altitude Adaptation

Poikilothermic animals living in high-altitude environments can be greatly affected by the anaerobic metabolism and lactate recycling, which are catalyzed by an enzyme called lactate dehydrogenase(LDH). However, the function and possible regulatory mechanisms of their anaerobic glycolysis remained elusive. We compared the difference in LDH between a native high-altitude(4 353 m) lizard, Phrynocephalus erythrurus, and a closely related species, Phrynocephalus przewalskii that lives in intermediate altitude environment(1 400 m). The activity of LDH, the concentration of lactate, the distribution of isoenzyme, and the mRNA amounts of Ldh-A and Ldh-B were determined. In cardiac muscle, the lactate-forming activity of P. erythrurus in LDH was higher than of P. przewalskii LDH at all three temperatures tested(10 °C, 25 °C and 35 °C), while lactate-oxidation activity of LDH was significantly different between the two species only at 25 °C and 35 °C. In skeletal muscle, both lactate-forming and lactate-oxidation rates of P. erythrurus were lower than that of P. przewalskii. There was a higher proportion of H subunit and a significantly higher expression of Ldh-B, with a concomitant decrease of lactate concentration in P. erythrurus. These results indicate that P. erythrurus may have a strong potential for anaerobic metabolism, which is likely adapted to the hypoxic environment at high altitudes. Furthermore, P. erythrurus is capable of oxidizing more lactate than P. przewalskii. The Ldh-A cDNA of the two species consists of a 999 bp open reading frame(ORF), which encodes 332 amino acids, while Ldh-B cDNA consists of a 1 002 bp ORF encoding 333 amino acids. LDHA has the same amino acid sequence between the two species, but three amino acid substitutions(V12 I, N21S and N318K) were observed in LDHB. Structure analysis of LDH indicated that the substitutions of residues Val12 and Asp21 in P. erythrurus could be responsible for the highaltitude adaptation. The LDH characteristics of LDH in P. erythrurus suggest unique adaptation strategies of anaerobic metabolism in hypoxia and cold environments at high altitudes for poikilothermic animals.

High serum lactate dehydrogenase and dyspnea:Positive predictors of adverse outcome in critical COVID-19 patients in Yichang

BACKGROUND Coronavirus disease 2019(COVID-19)caused by the severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)outbreak in China,constitutes a Public Health Emergency of International Concern.It is well known that COVID-19 patients may have increased serum lactate dehydrogenase(LDH)levels in the early stage.The clinical changes in LDH may have predictive value in disease evolution and prognosis in critically ill COVID-19 patients.AIM To examine serum LDH and clinical characteristics in patients with COVID-19 and their predictive value for prognosis.METHODS This retrospective study analyzed the clinical data of forty-seven critical COVID-19 patients in the intensive care unit of the Third People's Hospital of Yichang City from January 27 to March 25,2020 and divided them into survivors and nonsurvivors.The patients were diagnosed according to the World Health Organization interim guidance and critical cases met any one of the following criteria:Respiratory failure and required mechanical ventilation,the occurrence of shock,and the combined failure of other organs that required intensive care unit monitoring and treatments,according to the diagnostic criteria of critical COVID-19.Clinical data including symptoms,detection of SARS-CoV-2,chest computed tomography(CT)images,changes in serum LDH in different clinical phases,and prognosis were collected.Statistical analysis of the data was performed.Continuous variables were expressed as median(interquartile range)and compared with the Mann-Whitney U test.Categorical variables were compared with the Chi-square test.Survival data were analyzed using Kaplan-Meier survival curves and log-rank tests.RESULTS According to chest CT images,we observed the alveolitis and fibrosis stages in all critical patients in this study.Most non-survivors died in the fibrosis stage.Nonsurvivors had fewer days of hospitalization,shorter disease duration,shorter duration of alveolitis and fibrosis,and had dyspnea symptoms at disease onset(P=0.05).Both first and lowest LDH values in the alveolitis stage were more pronounced in non-survivors than in survivors(449.0 U/L vs 288.0 U/L,P=0.0243;445.0 U/L vs 288.0 U/L,P=0.0199,respectively),while the first,lowest and highest values of serum LDH in non-survivors were all significantly increased compared to survivors in the fibrosis phase(449.0 U/L vs 225.5 U/L,P=0.0028;432.0 U/L vs 191.0 U/L,P=0.0007;1303.0 U/L vs 263.5 U/L,P=0.0001,respectively).The cut-off points of first LDH values in the alveolitis and fibrosis phase for distinction of non-survivors from survivors were 397.0 U/L and 263.0 U/L,respectively.In the fibrosis stage,non-survivors had more days with high LDH than survivors(7.0 d vs 0.0 d,P=0.0002).Importantly,patients with high LDH had a significantly shorter median survival time than patients with low LDH in the alveolitis phase(22.0 d vs 36.5 d,P=0.0002),while patients with high LDH also had a significantly shorter median survival time than patients with low LDH in the fibrosis phase(27.5 d vs 40.0 d,P=0.0008).The proportion of non-survivors with detectable SARS-CoV-2 until death in the alveolitis stage was significantly increased compared with that in the fibrosis stage(100%vs 35.7%,P=0.0220).CONCLUSION High LDH and dyspnea symptoms were positive predictors of an adverse outcome in critical COVID-19.The rapid progressive fibrosis stage was more perilous than the alveolitis stage,even if SARS-CoV-2 is undetectable.

Predictive value of serum lactate dehydrogenase in diagnosis of septic shock in critical pediatric patients:A cross-sectional study

Objectives:To determine the predictive value of lactate dehydrogenase(LDH)in the diagnosis of septic shock and its association with other prognostic scores in critical pediatric patients.Methods:A cross-sectional study was performed at Children’s Hospital of Cairo University between June 2019 and December 2019.A total of 200 pediatric patients were divided into the septic shock group[100 critically ill patients with septic shock from the pediatric intensive care unit(PICU)]and the control group(100 patients with only sepsis).LDH was determined in the first 24 hours of admission.The sensitivity and specificity of LDH in diagnosis of septic shock were assessed;the levels of related indicators of patients with different etiologies were compared;correlations between LDH,Paediatric Index of MortalityⅡ,and Pediatric Sequential Organ Failure Assessment(pSOFA)were analyzed.Results:Median LDH was 512μL(406.50-663.00)in the septic shock group and was significantly higher than that[190μL(160.00-264.50)]in the control group(P<0.001).Besides,median LDH in children with chest infecion was higher than that in children with other diagnoses(P=0.047).A good positive correlation was found between pSOFA and LDH(r=0.503,P<0.001).Conclusions:LDH could be a potential inflammatory marker in the diagnosis of septic shock and is valuable for PICU admission decisions.

Differential pulse voltammetric studies on the effects of Al(Ⅲ) on the lactate dehydrogenase activity

In this paper, differential pulse voltammetry （DPV） was applied to study the effects of aluminum Al（Ⅲ） on the lactate dehydrogenase （LDH） activity. Michaelis-Menten constant （Km^NADH） and maximum velocity （vmax） in the enzyme promoting catalytic reaction of ＂pyruvate（Pyr） ＋ NADH ＋ H^＋ ↑LDH←→lactate ＋ NAD^＋＂ under different conditions by monitoring DPV reduction current of NAD^＋ were reported.

Increased levels of lactate dehydrogenase and hypertension are associated with severe illness of COVID-19

BACKGROUND Coronavirus disease 2019(COVID-19)is amid an ongoing pandemic.It has been shown that patients with cardiovascular comorbidities are at higher risk of severe illness of COVID-19.AIM To find out the relationship between cardiovascular comorbidities and severe illness of COVID-19.METHODS The clinical data of 140 COVID-19 patients treated from January 22,2020 to March 3,2020 at our hospital were retrospectively collected.The clinical characteristics were compared between patients with mild illness and those with severe illness.RESULTS There were 75 male patients and 65 female patients(53.6%vs 46.4%).The mean age was 45.4±14.6 years(range,2-85 years).Most of the patients had mild illness(n=114,81.4%)and 26 patients had severe illness(18.6%).The most common symptom was fever(n=110,78.6%),followed by cough(n=82,58.6%)and expectoration(n=51,36.4%).Eight patients were asymptomatic but were positive for severe acute respiratory syndrome coronavirus 2 RNA.Patients with severe illness were significantly more likely to be hypertensive than those with mild illness[(10/26,38.4%)vs(22/114,19.3%),P=0.036].The levels of lactate dehydrogenase were significantly higher in the severe illness group than in the mild illness group(299.35±68.82 vs 202.94±63.87,P<0.001).No patient died in either the severe illness or the mild illness group.CONCLUSION Hypertension and elevated levels of lactate dehydrogenase may be associated with severe illness of COVID-19.

Lactate Dehydrogenase Isoenzymes in Mammalian Cells Exposed to Isophenphos

The cytotoalcity of isophenphos, an organophosphorus insecticide that has the potential to cause delayed polyneuropathy, was evaluated in GBK and V79 ce1ls. A 72 h time course following isophenphos exposure indicated a dose-dependent growth inhibition as dctermined by cell counts. The administration of isophenphos (20g/ml) to GBK cells cultured at high densities indicated a decrease in the activities of LDH isocnzymes. Analysis of V79 cells revealed a decrease of LDH3, the only LDH isocnzyme detected in these cells.

Changes of lactate dehydrogenase in corneal edema after cataract surgery treated with trans-corneal oxygenation therapy

AIM:To investigate the changes in levels of the lactate dehydrogenase(LDH)enzyme in corneal edema after cataract surgery with trans-corneal oxygenation therapy.METHODS:This pre-post design study design conducted on 15 patients with corneal edema after cataract surgery and receiving trans-corneal oxygenation therapy.Tear sample(using Schirmer paper,from the inferior fornix of the conjunctiva)was carried out prior to trans-corneal oxygenation therapy,on the day 2(D2)and day 5(D5)postoperatively before and after trans-corneal oxygenation therapy.Visual acuity[VA(Log MAR)],corneal endothelial density,central corneal thickness(CCT),and coefficient of variation corneal endothelial(Co V)were recorded.The value of LDH was measured using ELISA.The difference in mean LDH value before and after trans-corneal oxygenation therapy,between two groups were analyzed using Wilcoxon signed rank test.RESULTS:There was a decrease in LDH tear concentration at D2(pre vs post:1127.54±497.09 vs 696.91±489.49;P=0.002)and D5(pre vs post:1064.17±677.77 vs 780.28±428.95;P=0.027)after trans-corneal oxygenation therapy as well as decrease in LDH concentration on the D2 compared to D5(P=0.041).The mean CCT was decreased significantly after the administration of trans-corneal oxygenation(pre vs post:632.10±25.66 vs 563.90±51.54;P=0.005).The mean VA and Co V increased significantly after the administration of trans-corneal oxygenation(P=0.001 and P=0.028,respectively).However,there was no difference in mean of corneal endothelial density(P=0.814).CONCLUSION:Trans-corneal oxygenation therapy is associated with significant decrease of tears LDH levels in post cataract surgery with corneal edema.It is accompanied by clinical improvement such as significant reduction of CCT.

Molecular Weight of Cytoplasmic Malate Dehydrogenase,Mitochondrial Malate Dehydrogenase and Lactate Dehydrogenase of a Freshwater Catfish

The multiple molecular forms of cytoplasmic malate dehydrogenase (cMDH), mitochondrial malate dehydrogenase (mMDH ) and lactate dehydrogenase (LDH ) were studied in the liver and skeletal muscle of the freshwater catfish, Clarias batrachus. There were two electrophoretically distinguishable bands (AA andBB) of cMDH and mMDH which suggests that they are apparently encoded at two gene loci (A and B) in both the tissues.However, the presence of a single band (LDH-1 ) of LDH in liver and double bands (LDH-1and LDH-2) in skeletal muscle in which LDH-2 was predominant reflects the differential expression of LDH genes in different metabolic tissues to meet the requirement of energy production. The AA isoform (74 kd) of liver cMDH was smaller than those of the AA form (110 kd) of skeletal muscle. In contrast, the BB isoform of liver (42 kd) and skeletal muscle (54 kd) were more or less similar in size. Unlike the case of cMDH, the molecular weight of AA isoform (115 kd) of liver mMDH was higher than those of the AA form (87kd) of skeletal muscle. Whereas the molecular weight of BB isoform (58 kd) of liver was in proximity to the weight of BB form (44 kd) of skeletal muscle mMDH. The size of AA isoform (74 kd) of liver cMDH was smaller, while the AA isoform (110 kd) of skeletal muscle was larger as compared to AA form of mMDH in the liver (115 kd) and skeletal muscle (87 kd). But the size of BB isoform of both the isozymes was almost equal in these metabolic tissues. The molecular weight of liver LDH-1 (96 kd) was close to the weight of LDH-1 (82 kd) in skeletal muscle. The molecular weight of skeletal muscle LDH-2 was deduced as 37 kd which is much more lower than the weight of LDH-1 in liver and skeletal muscle. The smaller size of LDH-2 in skeletal muscle may be of a physiological significance in this anaerobic tissue

Escherichia coli Expression of Lactate Dehydrogenase C Gene of Black-Lipped Pika in Western Sichuan Plateau

[ Objective] To express lactate dehydrogenase C （LDH-C） gene in prokaryotic system and then purify the recombinant protein. E Meth- od] The LDI-I-C gene was amplified from black-lipped pika（ Ochotona curzoniae） by RT-PCR and ligated to the expression vector pET-32a. The E. coli BL21 （DE3） carrying the LDI-I-C gene was obtained and induced by IPTG （isopropy-beta-D-thiogalactoside）. The expression products were analyzed by SDS-PAGE and purified by affinity chromatography. L Result] An approximately 1.0 kbp band appeared in the RT-PCR products as the- oretically expected. As evidenced by PCR and double enzyme digestion, an approximately 1.0 kbp fragment appeared, which indicated successful construction of expression vector. As analyzed by SDS-PAGE, a fusion protein with molecular weight slightly larger than 45 kDa was expressed in form of inclusion body. And very pure fusion protein was obtained by nickel ions-charged affinity chromatography. [ Coaclusion] The LDH-C gene of black-lipped pika has been cloned and expressed in E. coil

A Review on Molecular Physiology of Malate and Lactate Dehydrogenases in Fishes

The malate(EC 1.1.1.37)and lactate(EC 1.1.1.27)dehydrogenases are themetabolic enzymes directly or indirectly involved in energy production,gluconeogenesis and lipogenesis.Malate dehydrogenase(MDH)exists in twoisoenzymic forms,cytoplasmic(cMDH)and mitochondrial(mMDH),composed of Aand/or B subunits(dimeric molecule:MW 40,000-120,000).Lactate dehydrogenase(LDH)has tetrameric(MW 35,000-110,000)structure made up of either A and/or B,orC(C,E,F)subunits.They catalyze an ordered bisubstrate(substrate and coenzyme)

Testicular germ cell tumors type 2 have high RNA expression of LDHB,the gene for lactate dehydrogenase subunit B

This study analyzed RNA expression of genes for three serum tumor markers,alpha fetoprotein(AFP),human chorionic gonadotropin(hCG),and lactate dehydrogenase(LDH),in patients with testicular germ cell tumors(TGCT)type 2.The gene AFP encodes AFP,the gene for chorionic gonadotropin beta polypeptide 5(CGB5)encodes a major part of the specific beta subunit of hCG,and the genes for LDH subunit A(LDHA),LDH subunit B(LDHB),and LDH subunit C(LDHC)encode three different subunits of LDH.LDHB encodes the LDHB subunit present as a tetramer in LDH isoenzyme 1(LDH-1).We examined three datasets with 203 samples of normal testis tissue(NT)and TGCT type 2.Yolk sac tumor(YST)expressed RNA of AFP fourteen thousand times higher than seminoma(SE),embryonal carcinoma(EC),and teratoma(TER)combined(P=0.00015).In the second microarray,choriocarcinoma(CC)expressed RNA of CGB5 ten times higher than other histologic types of TGCT combined.EC expressed RNA of LDHB twice higher than SE,YST and TER combined(P=0.000041).EC expressed RNA of LDHB higher than that YST expressed RNA of AFP and that CC expressed RNA of CGB5.In conclusion,TGCT type 2 expressed RNA of LDHB markedly higher than the RNA of 23 other candidate genes for TGCT type 2.

Lovastatin changes activities of lactate dehydrogenase A and B genes in rat myocardial cells

Background Lactate dehydrogenase （LDH） is a crucial regulator of energy metabolism in many organs including the heart. Lovastatin is widely used in prevention and treatment of coronary heart disease and is a drug with substantial metabolic influences. Our study aimed to determine the activities of the lactate dehydrogenase A and B （LDHA and LDHB）genes following lovastatin treatment.Methods The rat myocardial cell line H9c2（2-1） in culture was exposed to 100 nmol/L lovastatin for 24 hours or for five days. The functions of the LDHA and LDHB genes were examined at the transcriptional （mRNA） level with quantitative real-time polymerase chain reaction （Q-RT-PCR）, and at the translational （protein） level with immunoblotting.Results When compared with control levels, the LDHA mRNA went up by （151.65±16.72）% （P=0.0132） after 24 hours and by （175.28±56.54）% （P=0.0366） after five days of lovastatin treatment. Although 24 hours of lovastatin treatment had no significant effects on LDHB mRNA levels, when the treatment was extended to five days, LDHB mRNA levels were significantly down-regulated to （63.65±15.21）% of control levels （P=0.0117）. After 24 hours of treatment with lovastatin,there were no significant changes in protein levels of either LDHA or LDHB. When treatment time was extended to five days, the protein levels of LDHA were up-regulated by （148.65±11.81）% （P=0.00969）, while the protein levels of LDHB were down-regulated to （64.91±5.47）% of control levels （P=0.0192）.Conclusions Lovastatin affects gene activities of LDHA and LDHB differently, which may reveal novel pharmacological effects of lovastatin.

Construction of sperm-specific lactate dehydrogenase DNA vaccine and experimental study of its immuno-contraceptive effect on mice

Lactate dehydrogenase C4 (LDHC4) is a key enzyme for sperm metabolism. It is distributed specifically in testis and is highly immunogenic. In this study, two DNA vaccines pVAX1-hLDHC and pVAX1-mLDHC were constructed by inserting coding sequences of human and mice LDHC4 into the eukaryotic ex-pression vector pVAX1. The production of LDHC4 specific antibodies was induced in the sera of vac-cinated mice and the reproductive tract secretions of vaccinated female mice through immunization by mucosal surface instillation. Furthermore, the antibody titer increased with the times of immunization. In the mating experiment, the number of newborns of the vaccinated mice reduced significantly and some immunized female mice even lost the ability to bear any offsprings, suggesting that the difference between the immunized and control mice was statistically significant. Sperm agglutination analysis indicated that both the antisera from immunized mice and the reproductive tract secretions of vacci-nated female mice could agglutinate normal sperms. Results of immunohistochemistry showed that the antibodies present in the sera of immunized mice and the reproductive tract secretions of vaccinated female mice could specifically react with LDHC4 antigen, which mainly locates in the cytoplasm, acrosome membrane externa and acrosome capsule of the sperm. Taken together, our results indicated that the constructed contraceptive DNA vaccines did yield immunocontraceptive effects on mice and this would enable clinical trials in near future.

Single amino-acid substitution in the N-terminal arm altered the tetramer stability of rat muscle lactate dehydrogenase A

Lactate dehydrogenase A (LDHA) is a well-characterized tetrameric enzyme. Its N-terminal arm, comprised of an (-helix and a (-strand, was suggested to be essential for subunit interactions. To examine the critical amino acid residues in the N-terminus involved in the subunit association, two single-point mutants, Leu3Pro (L3P) and Ile8Glu (I8E), have been constructed. We compared the stability of WT-LDHA (WT) and its variants by unfolding experiments. For WT, a dimeric but inactive intermediate was observed by size-exclusion chromatography at 0.6-0.8 mol/L GdmCl. Leu3Pro exists in an active tetrameric structure in aqueous solution as WT does, but it dissociates into dimers under lower concentration of GdmCl (0.2 mol/L). In aqueous solution, the Ile8Glu variant exists predominantly in the dimeric form with increased KM and decreased kcat as compared with those of WT and L3P. However, the activity of Ile8Glu increases significantly in the presence of sodium sulfate. In conclusion, two mutants are less stable than WT in oligomer structure. Results also support the fact that some residues in the N-terminal arm, especially the Leu8 in the (-structure, contribute the important binding energies to the dimerization of dimers, which might affect the assembly of the enzyme as well as the catalytic function.

Inactivation of Lactate Dehydrogenase from Pig Heart by o-Phthalaldehyde

Treatment of lactate dehydrogenase (LDH) with o phthalaldehyde resulted in a time dependent loss of enzyme activity. The inactivation followed pseudo first order kinetics over a wide range of the inhibitor. The second order rate constant for the inactivation of LDH was estimated to be 1.52 (mol/L) -1 ·s -1 . The modified enzyme showed a characteristic fluorescence emission spectrum with a maximum at 405 nm upon excitation at 337 nm, consistent with the formation of isoindole derivatives by the cross linking of proximal cysteine and lysine residues. The loss of enzyme activity was concomitant with the increase in absorbance at 337 nm. Stoichiometric study of the reaction showed that complete loss of activity was accompanied by formation of approximately four moles of isoindole derivatives per mole of LDH subunits. One of the substrates, NADH, partially prevented the enzyme from reacting with o phthalaldehyde, whereas the other substrate, pyruvate, did not provide any protection. Protection experiments suggest that one of the cysteine lysine pairs modified by o phthalaldehyde is near the NADH binding site of LDH.

DETECTION OF PLASMODIUM FALCIPARUM LACTATE DEHYDROGENASE A PROMISING APPROACH TO QUICK DIAGNOSIS OF MALARIA

To establish a quick and applicable diagnostic method has been a major target in immunological research on malaria. Of all approaches studied, circulating antigen (CAg) detection seems to be the most promising. Scientists have encountered tough difficulties in the analysis and purification of malaria parasite antigen because of its high complexity. Studies showed that plasmadium falciparum (P.f) lactate dehydrogenase (pLDH), as a specific CAg of malaria parasite. is apparently different from that of human erythrocytes (rLDH) both in their physical and biochemical characteristics. and is very easy to be identifed. Consequently, pLDH detection has a great potential to be developed into a method for assessing parasitemia. This paper reviewed the methods for the purification. separation, identification of pLDH and the prospect of its clinical applications as an ideal detector of the presence of malaria parasite in order to speed up this research.

Lysine acetylation decreases enzyme activity and protein level of Escherichia coli lactate dehydrogenase

Lactate is an important bulk chemical with widespread applications and a major byproduct of other chemicals bioprocess in microbial fermentation.Lactate dehydrogenase A(LdhA)catalyzes the synthesis of lactate from pyruvate.Lysine acetylation is an evolutionarily conserved post-translational modification;however,the mech-anisms underlying the regulation of LdhA function by lysine acetylation in Escherichia coli remain poorly under-stood.Herein,we demonstrate acetylation of E.coli LdhA occurs via enzymatic and non-enzymatic mechanisms.Further,we show carbon source type and concentration affect the lysine acetylation status of LdhA via a non-enzymatic mechanism.Lysine acetylation significantly inhibits the enzymatic activity and protein level of LdhA.The results of the present study demonstrate lysine acetylation of E.coli LdhA is irreversible.Understanding of the effects of lysine acetylation on LdhA function may provide a new perspective for regulating lactate production in microbial synthesis.