Isozyme Patterns Research Articles

SOME BIOCHEMICAL PROPERTIES OF CATALASE FROM SAFFLOWER (CARTHAMUS TINCTORIUS L.CV. IL-111)

Safflower (Carthamus tinctorius L.cv) is a member of the family Compositae that is cultivated mainly for its seed, which is used as edible oil and bird seed. Catalase is a common enzyme found in nearly all living organisms that are exposed to oxygen, where it functions to catalyse the decomposition of hydrogen peroxide to water and oxygen. In this study catalase was extracted from leaves of safflower (cv. M-CC-190) using phosphate buffer (0.1 M, pH 7.2) and its kinetic properties, thermal stability, isozymic pattern and sensitivity to inhibitors were investigated. On the basis of pH profile and activity staining for catalase on native PAGE, two isoform of catalases were detected in extract with pH optima at 6.5 and 8.5, and the range of catalase activity was found between 5.5 and 10.5. Apparent Km and Vmax of catalase at two pH optima were different, but the catalytic efficiency values were similar. Effect of heat treatment on catalase activity showed lower heat stability for isoenzyme active at pH 8.5. Isoenzyme with pH optima at 6.5 was more sensitive to both azide and cyanide in comparison to other isoenzyme active in M-CC-190 leaves extract. Key words: Catalase, safflower, isoenzyme, kinetic, thermal stability, M-CC-190.

Differential responses of the antioxidant defence system and ultrastructure in a salt-adapted potato cell line

Changes in lipid peroxidation and ion content and the possible involvement of the antioxidant system in salt tolerance at the cellular level was studied in a potato (Solanum tuberosum L.) callus line grown on 150 mM NaCl (salt-adapted) and in a non-adapted line exposed to 150 mM NaCl (salt-stressed). Salinity reduced the growth rate and increased lipid peroxidation in salt-stressed line, which remained unaltered in the adapted line. Na⁺ and Cl⁻ content increased due to salinity in both lines, but the adapted line displayed greater K⁺/Na⁺ ratio than the stressed one. Total superoxide dismutase (SOD, EC 1.15.1.1), ascorbate peroxidase (APX, EC 1.11.1.11), and glutathione reductase (GR, EC 1.6.4.2) activities decreased in both salt-exposed lines; catalase (CAT, EC 1.11.1.6) activity did not change in the adapted line, but decreased in the stressed cell line. Salinity caused the suppression of one GR isoform, while the isozyme patterns of SOD, APX, and CAT were not affected. Ascorbate and reduced glutathione increased in both salt-exposed calli lines. α-Tocopherol increased as a result of salt exposure, with higher levels found in adapted calli. Electron microscopy showed that neither the structural integrity of the cells nor membrane structure were affected by salinity, but plastids from adapted cells had higher starch content. The results suggest that the enzymic and non-enzymic components of the antioxidant system are differentially modulated by salt. Different concentrations of antioxidant metabolites are more relevant to the adaptive response to salinity in potato calli than the differences in activity of the antioxidant enzymes.

Modulation of endogenous peroxidase by exogenous peroxidase in chinese red radish seedling

To investigate the peroxidase function and its signal pathway, activity and isozyme pattern of peroxidase and transcription level of the peroxidase gene rsprx1 and rsprx2 in Chinese red radish seedling (Raphanus sativus L.) were investigated after administration of the exogenous peroxidase (extract of radish). The activity and isozyme pattern of peroxidases were increased, among cotyledon, hypocotyl and root, the most significant increase of peroxidases activity was observed in hypocotyls, the activity of phenylalanine ammonialyase was also increased, but the pelargonidin content was decreased. Although the exogenous application of horseradish peroxidase and the extract of radish could both affect the activity of endogenous enzyme and metabolites of Chinese red radish seedlings, the latter was more effective than the former. Nifedipine, as L-calcium channel blocker, affected the activity of peroxidase and pelargonidin content in Chinese red radish seedling. Exogenously added nifedipine with the extract caused reversal of the effects. It is concluded that calcium was involved in the signal pathway of the extract of radish in Chinese red radish seedling.

Isozymes of antioxidative enzymes during ripening and storage of ber (Ziziphus mauritiana Lamk.)

Isozyme profile of antioxidative enzymes viz. superoxide dismutase (SOD), peroxidase (POX), catalase (CAT) and ascorbate peroxidase (APX) was studied during ripening and storage of two cultivars of ber fruit (Ziziphus mauritiana Lamk.) differing in their shelf-lives viz. Umran (shelf-life, 8-9d) and Kaithali (shelf-life, 4-5d). The profile revealed that Umran variety exhibited three bands each of SOD and POX while in Kaithali, these enzymes had two isoenzymes throughout ripening. CAT and APX, however, showed two isozymes each during ripening of both the varieties and the pattern remained the same at all the stages of ripening except at the initial stage i.e immature green stage where single CAT isozyme was visible. During storage, one extra band each of SOD and POX present only in Umran got disappeared at later stages of storage, whereas in Kaithali, the pattern remained unchanged. Also, there was no change in the pattern of CAT and APX isozymes during storage of both the varieties. One isozyme of CAT could be considered as ripening related while one isozyme each of SOD and POX could be related to higher shelf life of fruits.

Enzymatic features of the glucose metabolism in tumor cells

Many tumor types exhibit an impaired Pasteur effect, i.e. despite the presence of oxygen, glucose is consumed at an extraordinarily high rate compared with the tissue from which they originate - the so-called 'Warburg effect'. Glucose has to serve as the source for a diverse array of cellular functions, including energy production, synthesis of nucleotides and lipids, membrane synthesis and generation of redox equivalents for antioxidative defense. Tumor cells acquire specific enzyme-regulatory mechanisms to direct the main flux of glucose carbons to those pathways most urgently required under challenging external conditions such as varying substrate availability, presence of anti-cancer drugs or different phases of the cell cycle. In this review we summarize the currently available information on tumor-specific expression, activity and kinetic properties of enzymes involved in the main pathways of glucose metabolism with due regard to the explanation of the regulatory basis and physiological significance of the Warburg effect. We conclude that, besides the expression level of the metabolic enzymes involved in the glucose metabolism of tumor cells, the unique tumor-specific pattern of isozymes and accompanying changes in the metabolic regulation below the translation level enable tumor cells to drain selfishly the blood glucose pool that non-transformed cells use as sparingly as possible.

Changes to the proteome and targeted metabolites of xylem sap in Brassica oleracea in response to salt stress

Root-to-shoot signalling via xylem sap is an important mechanism by which plants respond to stress. This signalling could be mediated by alteration in the concentrations of inorganic and/or organic molecules. The effect of salt stress on the contents of xylem sap in Brassica olarecea has been analysed by mass spectrometry in order to quantify these changes. Subcellular location of arabinogalactan proteins (AGPs) by immunogold labelling and peroxidase isozymes was also analysed by isoelectrofocusing. The xylem sap metabolome analysis demonstrated the presence of many organic compounds such as sugars, organic acids and amino acids. Of these, amino acid concentrations, particularly that of glutamine, the major amino acid in the sap, were substantially reduced by salt stress. The xylem sap proteome analysis demonstrated the accumulation of enzymes involved in xylem differentiation and lignification, such as cystein proteinases, acid peroxidases, and a putative hydroxycinnamoyl-CoA:shikimate hydroxycinnamoyl transferase under salt stress. The peroxidase isozyme pattern showed that salt stress induced a high accumulation of an acid isoform. These results suggest that xylem differentiation and lignification is induced by salt stress. The combination of different methods to analyse the xylem sap composition provides new insights into mechanisms in plant development and signalling under salt stress.

Some biochemical properties of guaiacol peroxidases as modified by salt stress in leaves of salt-tolerant and salt-sensitive safflower ( Carthamus tinctorius L.cv. ) cultivars

The kinetics properties of guaiacol peroxidase (GP) and its isozymic pattern, and lipid peroxidation product were comparatively analyzed in two varieties of safflower ( cv. M-CC-190 as salt-tolerant and cv. IL-111 as salt-sensitive cultivars) under normal and different concentrations of NaCl. The pH profile of GP activity in leaves extract of two cultivars in control and salt stressed plants showed different pattern of pH dependency with three maxima peaks at pH 4.5, 6.5 and 8 in salt-tolerant cultivar and two maxima peaks at pH 4.5 and 6.5 in salt-sensitive cultivar. Comparison of catalytic efficiency for GP between two cultivars at respective pH, showed that, salt-tolerant cultivar in both control and salt stressed condition had higher catalytic efficiency than salt-susceptible cultivar. The GP activity on the gels revealed four and two isoforms of peroxidases in salt-tolerant and salt-sensitive cultivars, respectively. GPs increased their expression with higher levels of salinity. However, in salt-sensitive cultivar GPs expression exhibited threshold behavior, with increase expressions in isoenzymes up to a certain level of salinity (25 mM NaCl), followed by decrease to a level of expressions corresponding to the control groups. The levels of lipid peroxidation as indicated by thiobarbituric acid reactive substances (TBARS) were higher in the sensitive variety than the tolerant under control and NaCl salinity. The overall results obtained in this study suggest that, oxidative stress may play an important role in salt-stressed safflower plants and that the greater protection of M-CC-190 leaves from salt-induced oxidative damage results, at least in part, through the increase of the GPs activity, catalytic efficiency and induction of specific isoenzymes (P1 and P4).

Changes in fungal population of fly ash and vinasse mixture during vermicomposting by Eudrilus eugeniae and Eisenia fetida: Documentation of cellulase isozymes in vermicompost

Fly ash (FA) and vinasse (VN), two industrial wastes, are generated in huge amounts and cause serious hazards to the environment. In this experiment, different proportions of these two wastes were used as food for two epigeic earthworms ( Eisenia fetida and Eudrilus eugeniae) to standardize the recycling technique of these two wastes and to study their effect on fungal especially cellulolytic fungal population, cellulase activity and their isozyme pattern, chitin content and microbial biomass of waste mixture during vermicomposting. Increasing VN proportion from 25% to 50% or even higher, counts of both fungi and cellulolytic fungi in waste mixtures were significantly ( P ⩽ 0.05) increased during vermicomposting. Higher cellulase activity in treatments having 50% or more vinasse might be attributed to the significantly ( P ⩽ 0.05) higher concentration of group I isozyme while concentrations of other isozymes (group II and III) of cellulase were statistically at par. Higher chitin content in vinasse-enriched treatments suggested that fungal biomass and fungi-to-microbial biomass ratio in these treatments were also increased due to vermicomposting. Results revealed that Eudrilus eugeniae and Eisenia fetida had comparable effect on FA and VN mixture during vermicomposting. Periodical analysis of above-mentioned biochemical and microbial properties and nutrient content of final vermicompost samples indicated that equal proportion (1:1, w/w) of FA and VN is probably the optimum composition to obtain best quality vermicompost.

Genetic diversity in mulberry (Morus spp.) revealed by isozyme markers

SummaryNine isozyme patterns were studied to deduce the level of genetic diversity and inter-relationships among 14 species of mulberry (Morus spp. L) used in breeding programmes. βEsterase (EST), peroxidase (PoX), diaphorase (DIA), and polyphenol oxidase (PPO) produced 12 isozyme loci and a total of 22 alleles. The percentage of polymorphic loci and the average number of alleles per locus ranged from 50 – 75% and 1.336 – 1.667 respectively. Among 80 wild accessions of two wild and two domesticated species of Morus, 22 accessions of M. alba exhibited a higher number of polymorphic loci (75%), a higher polymorphic index (0.398), average number of alleles per locus (1.457), and expected heterozygosity (0.339) compared to the three other Morus species under study. Evidence for a higher rate of gene flow was found between populations of M. alba and M. indica compared to that between the other three species sampled. Phylogenetic clustering of the 80 wild mulberry accessions indicated three major groups. Accessions of the domesticated species grouped into one cluster, while accessions of the wild species grouped into one major and one minor cluster. Among all the isozyme loci, EST-2 was present only in polyploid species. The resolving power of isozymes was less than that of molecular markers, hence molecular markers should be used for phylogenetic studies in mulberry.

Capsicum Use in Cambodia: The Continental Region of Southeast Asia Is Not Related to the Dispersal Route of C. frutescens in the Ryukyu Islands

CapsicumUse in Cambodia: The Continental Region of Southeast Asia Is Not Related to the Dispersal Route ofC. frutescensin the Ryukyu Islands. The local nomenclature and use of Capsicum by Khmer and other ethnic groups in Cambodia and the distribution of the diagnostic ShDH-B isozyme pattern of C. frutescens were studied. People in Cambodia use Capsicum in various ways, not only as a condiment but also as a vegetable, a medicine, and a colorant, and in popular beliefs, agricultural rituals, taboos, and rice malt. The findings showed that the ShDH-B phenotype may not have occurred as a mutation in Asia but in the Americas and then was introduced to Asia. Also, the ShDH-B phenotype is distributed in the insular regions of Southeast and East Asia and Oceania, but seems not to be distributed in the continental region of Southeast Asia. One possible hypothesis is that C. frutescens possessing the ShDH-B phenotype was introduced directly from the Americas via Oceania to the Philippines, and it thereafter dispersed into the insular regions.

Response of Antioxidative Enzymes to Cadmium Stress in Leaves and Roots of Radish (Raphanus sativus L.)

Presented study has demonstrated that exposure of plants to toxic heavy metal Cd results a reduction in plant growth. Varied concentrations of CdCl2, ranging from 0.0 to 50 ppm in the germinating media reduced leaf area of radish plant, chlorophyll and carotenoid contents. Greater loss of chlorophyll b content than chlorophyll a was observed especially under 50 ppm Cd exposure. With regards to the distribution of Cd in roots and leaves, the obtained data showed that the maximum accumulation of Cd occurred in roots followed by leaves. Generally, Fe, Zn, Mn and Cu declined in leaves compared to the roots. Furthermore, substantial increases were observed in antioxidant enzymes, such as catalase (CAT), glutathione S-transferase (GST) and peroxidase (POD), in Cd-stressed plants in comparison with control. The Cd stress also induced several changes in CAT and POD isozyme profiles and enhanced their activities. The results suggest that the reduction of leaf area and pigment content together with antioxidant enzymes and isozyme patterns can be used as indicators to Cd contamination.

Expression Patterns of PR proteins with Different Extract Methods during Germination of Rape Seed (Brassica napus L.)

This study was conducted to investigate the expression patterns of pathogenesis-related proteins (chitinase, β-1,3-glucanase and peroxidase) using activity staining of native-polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate (SDS)-PAGE during germination of rape seed (Brassica napus L. cv. Saturnin). The crude enzymes were extracted by distilled water (DW, pH 6.0) and 100 mM K-PO4 buffer (pH 7.0). The expression patterns of chitinase isozymes changed clearly on 10% native-PAGE gel with DW and K-PO4 buffer extract and on 12% SDS-PAGE gel with K-PO4 buffer extract, except for 12% SDS-PAGE conducted using DW during germination. The active bands of the chitinase isozymes were observed as four major bands (ch1, ch2, 86, and 78 kDa) and three minor bands (71, 60, and 54 kDa) on 10% native-PAGE gel conducted using DW and K-PO4 buffer extract. The two active bands on the 12% (w/v) SDS-PAGE gel presented as 34 and 29 kDa with DW extract, whereas one active band of 34 kDa was observed when the K-PO4 buffer extract was used. Active bands of β-1,3-glucanase isozymes changed slightly on 10% native-PAGE gel with DW and K-PO4 buffer extract during germination. The active band of β-1,3-glucanase isozymes were shown to have a high molecular weight (G1 and G2) on native-PAGE gel with DW extract at 0, 1, 2, and 3 days after germination, but not at 4 and 5 days. One active band of β-1,3-glucanase presented as G1 in the K-PO4 buffer extract. Active staining of peroxidase was stronger earlier in the DW extract than K-PO4 buffer extract at 2 days. The active bands showed as P1 and P2 in both DW and K-PO4 buffer extract at 5 days after germination.

In vitro digestion of protein sources by crude enzyme extracts of the spiny lobster Panulirus argus (Latreille, 1804) hepatopancreas with different trypsin isoenzyme patterns

The development of cost-effective and nutritionally adequate formulated diets is a key step in the sustainable expansion of spiny lobster aquaculture. Despite proteins are the major and most expensive component of diets, few studies are available on protein digestibility in spiny lobsters and such assessments have never been performed by in vitro methods. Two techniques were used for studying in vitro protein digestion of some common aquafeed ingredients by the spiny lobster Panulirus argus: i) the digestion cell in which the digestion products are removed by dialysis, and ii) electrophoresis which allows the visualization of the different protein fractions in the tested meals. Since three main trypsin isozyme patterns or phenotypes have been recently described in P. argus, the potential differences in protein digestion between individuals with different trypsin isozyme patterns were assessed. Results herein presented demonstrate for the first time in a crustacean species that the different trypsin phenotypes differ in protein digestion efficiency. Also, the digestion cell method was applied for the first time to a crustacea, proving to be sensitive to small changes in digestion efficiency. This method could be used in further in vitro studies for examining other aspects of spiny lobsters digestive process.

Transfer of transformed Lesquerella fendleri (Gray) Wats. chloroplasts into Orychophragmus violaceus (L.) O.E. Schulz by protoplast fusion

Asymmetric intergeneric hybrid plants were obtained through protoplast fusion between Orychophragmus violaceus (L.) O.E. Schulz and Lesquerella fendleri (Gray) Wats. The latter carried chloroplasts transformed with the fused aadA16gfp gene construct, conferring streptomycin–spectinomycin resistance and UV-induced green fluorescence. The somatic hybrids were selected using the properties of spectinomycin-induced plastid defects in “albino” O. violaceus plants (chloroplast recipient) combined with the γ-irradiation-induced inactivation of nuclei in plastid donor L. fendleri. The morphology and esterase isozyme pattern of the hybrid plant as well as the results of the PCR analysis of internal transcribed spacer of nuclear ribosomal DNA proved that the regenerated hybrids carried O. violaceus nuclei, while PCR amplification of the atpB–rbcL spacer and aadA16gfp gene fragments confirmed the presence of the transformed L. fendleri chloroplasts in these plants. Expression of the fused aadA16gfp gene construct was confirmed by sodium dodecylsulfate–polyacrylamide gel electrophoresis analysis and the resistance of the obtained plants to both streptomycin and spectinomycin.

Isozyme and PCR-based genotyping of epidemic Phytophthora colocasiae associated with taro leaf blight

The Oomycetous fungus Phytophthora colocasiae causing leaf blight of taro is widely distributed in India. Wide geographic range or sexual recombination provides genetic differentiation within this species. To determine how genetic variation is partitioned in P. colocasiae, 14 isolates were isolated from different regions of India, where the incidence of leaf blight is great. Molecular and biochemical techniques were employed for assessing and exploiting the genetic variability among isolates of P. colocasiae. Seven polymorphic enzyme systems revealed 23 isozyme patterns, each uniquely characterised by the presence or absence of electromorphs. Further, 10 oligodeoxynucleotide primers were selected for random amplified polymorphic DNA (RAPD) assays, which resulted in 123 polymorphic bands for 10 isolates of P. colocasiae. The data were entered into a binary matrix and a similarity matrix was constructed using a DICE similarity (SD) index. A UPGMA cluster based on SD values was generated using a NTSYS computer program. Shannon's index was used to partition genetic diversity. Similarly, isozymes and RAPDs yielded high estimates of genetic variability. Genetic diversity estimates via isozyme and RAPD pattern indicated 78.26% and 100%, respectively, total diversity among populations. This type of genetic variation in P. colocasiae indicates that variation due to asexual and/or possibly infrequent sexual mechanisms is possible and that genetic differentiation has taken place as a result of geographic isolation. The presence of larger than expected RAPD variation in isolates of P. colocasiae and the presence of distinct different zymotypes among these isolates suggests that genetic recombination (or less likely hybridisation) is at least possible in this fungus and that geographic differentiation has taken place. Even isolates obtained from the same habitat have different RAPD patterns, indicating that many populations of this fungus are made up of more than one genet and that few are derived clonally.

Deoxyribonucleases (DNases) in the cortex and endosome from the marine sponge Tethya aurantium

The presence and activity of deoxyribonucleases in the cortex and endosome sections from a sponge, the sea orange Tethya aurantium, were investigated. The maximal enzyme activity in sponge homogenate was detected at pH 4.27, pH 7.0 and pH 8.5–8.75. Among different specimens, several distinct patterns of neutral DNase isozymes were observed in the cortex section. In each investigated specimen the highest neutral DNase activity belonged to high molecular weight proteins (up to75 kDa). The acid DNases showed a low level of enzyme activity. In the endosome section the acid DNase activity was up to ten times higher than in the cortex and the presence of DNase II-like protein was detected. Neutral DNase, which expressed the highest enzyme activity in all the investigated specimens, has a molecular weight of 20 kDa and belongs to the DNase I-like family. The results indicate that the activity of neutral and acid DNases is related to sponge sections and their biological functions. The cortex, as the sponge section that communicates with the environment, expresses high interindividual variability and heterogeneity of neutral DNases, while the endosome section, where the intracellular digestion is localized, is a site of high acid DNase activity.

Sustainable effects of diatomite on the growth criteria and phytochemical contents of Vicia faba plants

Recent scientific findings proved that silicon feeding will provide benefits to plant growth and yield depending on the source of Si, the mode of application , the concentration used, the plant species , the mode of action and the biotic or abiotic factor involved. To better understand the intrinsic properties of Si , this work was conducted using a series of diatomite (diatomaceous earth)at rates of 0,2.5, 5 and 10 g/kg soil, to look into the most subtle changes in the growth , yield, metabolism, electropharetic protein and isozyme pattern of faba bean ( Vicia faba) plant. Indeed, diatomite-treated plants displayed an over all better morphological increments expressed as shoot and root length , number of leaves and pods/ plant, fresh and dry weights of shoots and roots and physiological activity expressed as chlorophyll a,b and total pigments , total soluble sugars (TSS) and total sugars (TS), photosynthetic rate, leaf stomatal conductance, % of leaf relative humidity ( %LRH), net intercellular carbon dioxide (∆CO2), total nitrogen (TN) and total soluble nitrogen (TSN) and the concentrations of phosphorus (P), potassium (K), calcium(Ca) and magnesium (Mg) than non-treated faba bean plants . Furthermore, diatomites (86-89 % SiO2) induced and restored ten electrophoretic protein bands which confirmed their significance as an algal manure in accelarating the gene functions to perform, in an elevated rate, new silence genes to operate thus expressing new protein profiles. Soil fertilization by diatomites induced four esterase ( EST), three peroxidase (POD), two catalase (CAT) and three acid phosphatase (ACP) isozyme polymorphic bands which were absent in the control plants .The intensity and density of these bands were significantly increased in response to diatomite treatment. The impact of diatomite is time-dependent as it became more effective while the experiment continued and increased in a dose-related manner . Reversibly, diatomites application showed negative response in the transpiration rate and the values of both carotenoids, sodium and iron. This study highlighted the fact that diatomites are important component of the production system and should not be ignored when attempting to attribute causes for below optimum production

Characterization of alcohol dehydrogenase 1 of the thermotolerant methylotrophic yeast Hansenula polymorpha

The thermotolerant methylotrophic yeast Hansenula polymorpha has recently been gaining interest as a promising host for bioethanol production due to its ability to ferment xylose, glucose, and cellobiose at elevated temperatures up to 48 degrees C. In this study, we identified and characterized alcohol dehydrogenase 1 of H. polymorpha (HpADH1). HpADH1 seems to be a cytoplasmic protein since no N-terminal mitochondrial targeting extension was detected. Compared to the ADHs of other yeasts, recombinant HpADH1 overexpressed in Escherichia coli exhibited much higher catalytic efficiency for ethanol oxidation along with similar levels of acetaldehyde reduction. HpADH1 showed broad substrate specificity for alcohol oxidation but had an apparent preference for medium chain length alcohols. Both ADH isozyme pattern analysis and ADH activity assay indicated that ADH1 is the major ADH in H. polymorpha DL-1. Moreover, an HpADH1-deleted mutant strain produced less ethanol in glucose or glycerol media compared to wild-type. Interestingly, when the ADH1 mutant was complemented with an HpADH1 expression cassette, the resulting strain produced significantly increased amounts of ethanol compared to wild-type, up to 36.7 g l(-1). Taken together, our results suggest that optimization of ADH1 expression would be an ideal method for developing H. polymorpha into an efficient bioethanol production strain.

Growth and isozyme pattern of seedlings from polyembryonic seed in the mulberry, Morus spp

Growth and isozyme pattern of seedlings from polyembryonic seed in the mulberry, Morus spp

Unique expression pattern of human nicotinamide mononucleotide adenylyltransferase isozymes in red blood cells

Nicotinamide mononucleotide adenylyltransferase (NMNAT) catalyzes the formation of nicotinamide adenine dinucleotide (NAD). In humans, three isozymes have been identified: NMNAT1, which is widely expressed in all tissues, NMNAT2 and NMNAT3, which show a tissue-specific expression and whose mRNA levels are generally lower compared to NMNAT1. In the present study we determined the individual NMNAT isozymes activity in human red blood cells (RBCs) by using a biochemical discrimination assay based on the distinctive catalytic properties of the three proteins. We found that isozyme 3 predominates over isozyme 1, whereas isozyme 2 is absent. This high prevalence of NMNAT3 is cell-aging independent and was also confirmed by analyzing the mRNA and protein levels. RBC represent the first human cell type with a remarkable predominance of NMNAT3, and this unique expression pattern is discussed in light of the catalytic properties of the isozymes and in consideration of the biochemical microenvironment of RBC.