Oxidative demethylation of t-butyl alcohol by rat liver microsomes

Tertiary butyl alcohol has often been used experimentally as a “non-metabolizable” alcohol. In this report, evidence is presented that t-butanol serves as a substrate for rat liver microsomes and that it is oxidatively demethylated to yield formaldehyde. The apparent K_m for t-butanol is 30 mM while V_max is about 5.5 nmol per min per mg microsomal protein. Formaldehyde production is stimulated by azide, which prevents destruction of H₂O₂ by catalase. Hydroxyl radical scavenging agents, such as benzoate, mannitol, and 2-keto-4-thiomethylbutyrate, suppress formaldehyde production. Therefore, the microsomal reaction pathway appears to involve the interaction of t-butanol with hydroxyl radicals generated from H₂O₂ by the microsomes. Formaldehyde is also produced when t-butanol is incubated with model hydroxyl radical-generating systems such as the iron-EDTA-stimulated oxidation of xanthine by xanthine oxidase or the iron-EDTA-catalyzed autoxidation of ascorbate. These results indicate that t-butanol cannot be used to distinguish metabolically-linked from non-metabolically-linked actions of ethanol.     

Cosolvent-buffer mixtures as models for the cytoplasmic milieu: The enzymology of adenosine aminohydrolase.

Summary Adenosine aminohydrolase from calf intestinal mucosa is sensitive to changes in its environment produced by small mole fractions of dimethylsulfoxide (DMSO). At a mole fraction of 0.1 where the dielectric constant is lowered from that of 78 of neat water to about 76.5,V _max was reduced by 65% and affinity for substrate (adenosine) and the two competitive inhibitors, inosine and N⁶-benzyladenosine, was decreased markedly. However, this decreased affinity was such that K_i/K_m remained virtually constant for both inhibitors. DMSO itself showed the kinetics of a mixed inhibitor with K_i decreasing with increasing mole fraction. This cosolvent also decreased the heat stability of the enzyme which suggests that enzyme conformation is altered by DMSO.Comparison of data in the presence of DMSO with previously obtained data with dioxane shows that heat stability as well asV _max, at a given value of dielectric constant, is independent of the amount or nature of cosolvent used to achieve that dielectric constant. However, cosolvent induced changes in Ki indicate that colligative as well as dielectric constant effects contribute to the observed changes in kinetic behavior.These experiments may be considered as models for the behavior of enzymes in the medium of lowered dielectric constant expected in the vicinity of cytoplasmic membranes. The results indicate that in such an environment, adenosine aminohydrolase would be expected to be less efficient a catalyst, but equally susceptible to product inhibition, as compared to media of dielectric constant approaching that of water.Supported in part by Grant RR-262 from the General Clinical Research Centers Progam of the Division of Research Resources, National Institutes of Health.     

Calponin-like protein from mussel smooth muscle is a competitive inhibitor of actomyosin ATPase

The goal of this work was to elucidate the mechanism of inhibition of the actin-activated ATPase of myosin subfragment-1 (S1) by the calponin-like protein from mussel bivalve muscle. The calponin-like protein (Cap) is a 40-kDa actin-binding protein from the bivalve muscle of the mussel Crenomytilus grayanus. Kinetic parameters V_max and K_ATPase of actomyosin ATPase in the absence and the presence of Cap were determined to investigate the mechanism of inhibition. It was found that Cap mainly causes increase in K_ATPase value and to a lesser extent the decrease in V_max, which indicates that it is most likely a competitive inhibitor of actomyosin ATPase. Analysis of V_max and K_ATPase parameters in the presence of tropomyosin revealed that the latter is a noncompetitive inhibitor of the actomyosin ATPase.     

Crown ether activates cholesterol oxidase in low water media

Kinetic studies of cholesterol oxidase-catalysed oxidation of cholesterol in water/2-propanol mixtures showed a decrease of V _max/K _m values on the increase of concentration of the organic co-solvent. Addition of 18-crown-6 to the reaction medium results in an increase of V _max up to 16 times, and V _max/K _m up to 8.4 times, enhancing the activity of cholesterol oxidase in 2-propanol/water (88:12 v/v) to 3.5 times compared to the level observed in 46% 2-propanol.     

Highly efficient enzymatic synthesis of an ascorbyl unstaturated fatty acid ester with ecofriendly biomass‐derived 2‐methyltetrahydrofuran as cosolvent

Enzymatic synthesis of ascorbyl undecylenate, an unsaturated fatty acid ester of ascorbic acid, was reported with biomass‐derived 2‐methyltetrahydrofuran (MeTHF) as the cosolvent. Of the immobilized lipases tested, Candida antarctica lipase B (CAL‐B) showed the highest activity for enzymatic synthesis of ascorbyl undecylenate. Effect of reaction media on the enzymatic reaction was studied. The cosolvent mixture, t‐butanol‐MeTHF (1:4, v/v) proved to be the optimal medium, in which not only ascorbic acid had moderate solubility, but also CAL‐B showed a high activity, thus addressing the major problem of the solvent conflict for dissolving substrate and keeping satisfactory enzyme activity. In addition, the enzyme was much more stable in MeTHF and t‐butanol‐MeTHF (1:4) than in previously widely used organic solvents, t‐butanol, 2‐methyl‐2‐butanol, and acetone. The much higher initial reaction rate in this cosolvent mixture may be rationalized by the much lower apparent activation energy of this enzymatic reaction (26.6 vs. 38.1–39.1 kJ/mol) and higher enzyme catalytic efficiency (V_max/K_m, 8.4 vs. 1.3–1.4 h^?1). Ascorbyl undecylenate was obtained with the yields of 84–89% and 6‐regioselectivity of >99% in t‐butanol‐MeTHF (1:4) at supersaturated substrate concentrations (60 and 100 mM) after 5–8 h. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 30:1005–1011, 2014     

Steady-state kinetic studies on the actin activation of skeletal muscle heavy meromyosin subfragments: Effects of skeletal,smooth and non-muscle tropomyosins

The addition of either smooth muscle or brain tropomyosin to skeletal muscle actoheavy meromyosin (HMM) or acto-myosin subfragment-1 (SF1) produces an activation of the actin-activated ATPase activity up to 100%. This contrasts with the opposite, inhibitory effect produced by skeletal muscle tropomyosin. The degree of activation or inhibition depends on the ionic conditions, which influence the affinities of tropomyosin and HMM or SF1 for actin as well as on the molar ratio of actin to myosin.Enzyme kinetic analysis indicates that the inhibitory effect of skeletal muscle tropomyosin results from an approximately six- to tenfold increase in the apparent affinity (K_app) of the myosin head for the F-actin-tropomyosin complex with a concomitant six- to tenfold reduction in the maximal turnover rate (V_max). Thus, there is no direct competition of skeletal muscle tropomyosin and myosin for the same site on actin. Brain tropomyosin has an opposite effect, decreasing the apparent affinity with concomitant increase in the V_max.The effect of smooth muscle tropomyosin is more complex. At high ratios of myosin to actin this tropomyosin produces the same change in the K_app as skeletal muscle tropomyosin but yields a value of V_max that is about twofold higher. At lower molar ratios (below about 1 to 5 myosin subfragments to actin) the activating effect of this tropomyosin remains unchanged while the apparent affinity decreases to that observed for pure F-actin.On the basis of these data as well as from experiments carried out at fixed actin and varying SF1 concentrations, it is concluded that tropomyosins act in general as allosteric un-competitive inhibitors or activators of actomyosin by increasing or reducing the co-operative activation of myosin by actin at the level of product release.     

β-Glucosidase activity is involved in scent production in Narcissus flowers

Extrinsic environmental cues and intrinsic developmental stages of the flower control the production of scent from flowers. Flowers emit scent only when they are open; yet, the precursors for the aromatic compounds are also present in buds, stored as non-fragrant glycosides in the vacuole. We demonstrate that in Narcissus flowers scent emission is concurrent with an increase in the activity of β-glucosidase. The inhibition in vivo of β-glucosidase activity decreases scent emission from Narcissus flowers. The β-glucosidase activity was partially purified and the K_m, V_max and inhibition by gluconic acid lactone was determined.     

Control analysis of photosynthate partitioning

Experiments were carried out to investigate the contribution of ADP-glucose pyrophosphorylase and the plastid phosphoglucosemutase to the control of starch synthesis. Mutants ofArabidopsis thaliana (L.) Heyhn. were constructed with 50% and 7% of the wild-type adenosine 5′-diphosphoglucose pyrophosphorylase (ADPGlc-PPase), or 50% and null plastid phosphoglucomutase (PGM). The changes in the steady-state rates of sucrose synthesis, starch synthesis and CO₂ fixation were measured in saturating CO₂ in low (75 μmol·m⁻²·s⁻¹) and high (600 μmol·m⁻²·s⁻¹) irradiance. In low irradiance, a 50% decrease of PGM had no significant effect on fluxes, while a 50% and 93% decrease of ADPGlc-PPase led to a 23% and 74% inhibition of starch synthesis. Decreased ADPGlc-PPase led to an increase of hexose phosphates, triose phosphates and fructose-1,6-bisphosphate. Fixation of CO₂ was not inhibited because the inhibition of starch synthesis was matched by a stimulation of sucrose synthesis. In high irradiance, a 50% decrease of PGM led to a 20% inhibition of starch synthesis. A 50% and 93% decrease of ADPGlc-PPase led to a 39% and 90% inhibition of starch synthesis. Sucrose synthesis was also inhibited, and the rate of photosynthesis was decreased. Decreased ADPGlc-PPase led to an increase of hexose phosphates but triose phosphates and fructose-1,6-bisphosphate did not increase. These results are used to estimate flux-control coefficients for these enzymes for starch synthesis. Firstly, the flux to starch is only controlled by ADPGlc-PPase in low irradiance, but control is redistributed to other enzymes in the pathway when a rapid flux is imposed, e.g. in high irradiance and CO₂. Secondly, reducing the rate of starch synthesis by decreasing the activity of enzymes in this pathway does not always lead to a compensating increase in the rate of sucrose synthesis. Thirdly, decreasing the activity of an enzyme by a factor of two compared to the remainder of the pathway often leads to it exerting very considerable control. Fourthly, each enzyme starts to exert considerable control when only a fraction of its V_max activity is being utilised in vivo, for example the maximum flux at ADPGlc-PPase never exceeded 20% of the V_max activity. The summation theory is also applied to check whether additional major control sites are required. In low irradiance, the efficiency of light harvesting will exert considerable control over the rate of starch synthesis.     

Hepatic hexose transport and the effect of N2-induced anoxia and KCN on this process

Hepatic hexose transport was characterized using 3-O-methyl-D-glucose, which is not metabolized by the liver. The kinetic parameters determined in the starved state were taken as basal values for the transport system which showed saturation kinetics with high V_max and K_m values of 161 nmol/mg dry wt./rnin and 39 mM respectively. In the fed state, the V_max was found to be increased nearly two-fold; this may be due to a phenomenon known as trans-stirnulation. The effects of N₂-induced anoxia and of KCN were investigated. In the fasted state, anoxia caused the transport characteristics V_max and K_m to decrease nearly two-fold whereas KCN had the opposite effect as the V_max and K_m were increased by three- and two-fold respectively. In the fed state, anoxia and KCN caused a marked decrease in the transport characteristics.     

Diurnal regulation of phosphoenolpyruvate carboxylase from crassula

Phosphoenolpyruvate carboxylase appears to be located in or associated with the chloroplasts of Crassula. As has been found with this enzyme in other CAM plants, a crude extract of leaves gathered during darkness and rapidly assayed for phosphoenolpyruvate carboxylase (PEPc) activity is relatively insensitive to inhibition by malate. After illumination begins, the PEPc activity becomes progressively more sensitive to malate. This enzyme also shows a diurnal change in activation by glucose-6-phosphate, with the enzyme from dark leaves more strongly activated than that from leaves in the light.

When the enzyme is partially purified in the presence of malate, the characteristic sensitivity of the day leaf enzyme is largely retained. Partial purification of the enzyme from dark leaves results in a small increase in sensitivity to malate inhibition.

Partially purified enzyme is found by polyacrylamide gel electrophoresis analysis to have two bands of PEPc activity. In enzymes from dark leaves, the slower moving band predominates, but in the light, the faster moving band is preponderant. Both of these bands are shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis to be composed of the same subunit of 103,000 daltons.

The enzyme partially purified from night leaves has a pH optimum of 5.6, and is relatively insensitive to malate inhibition over the range from pH 4.5 to 8. The enzyme from day leaves has a pH optimum of 6.6 and is strongly inhibited by malate at pH values below 7, but becomes insensitive at higher pH values.

Gel filtration of partially purified PEPc showed two activity peaks, one corresponding approximately to a dimer of the single subunit, and the other twice as large. The larger protein was relatively insensitive to malate inhibition, the smaller was strongly inhibited by malate.

Kinetic studies showed that malate is a mixed type inhibitor of the sensitive, day, enzyme, increasing K_m for phosphoenolpyruvate and reducing V_max. With the insensitive, night, enzyme, malate is a K type inhibitor, reducing the K_m for phosphoenolpyruvate, but having little effect on V_max. The inhibition of the insensitive enzyme by malate appears to be hysteretic, taking several minutes to be expressed during assay, probably indicating a change in the conformation or aggregation state of the enzyme.

Activation by glucose-6-phosphate is of the mixed type for the day form of the enzyme, causing both a decreased K_m for phosphoenolpyruvate and an increased V_max, but the night, or insensitive, form shows only an increase in V_max in response to glucose-6-phosphate.

     

A study of the substrate specificity of aminopeptidase N from Lactococcus lactis subsp. cremoris Wg2

A systematic study was made of the ability of aminopeptidase N from Lactococcus lactis subsp. cremoris Wg2 to hydrolyse different peptide substrates. The enzyme showed a marked preference for substrates containing arginine as the N-terminal residue but, to a lesser extent, was also capable of cleaving other residues such as lysine and leucine. There was a tendency for the activity to increase with the hydrophobicity index of the C-terminal residue of dipeptide substrates. It was also observed that the enzyme tended to have higher affinities but lower V _max values for tripeptides with hydrophobic C-terminal residues. The values determined for K _m and V _max increased with chain length for oligopeptides of the general formula Lys-Phe-(Gly) _n , the optimum, as determined from V _max/K _m, being when n = 4. Typical K _m values for the most effective substrates were in the range 0.2–0.6 mM.     

Sucrose translocation and storage in the sugar beet

Several physiological processes were studied during sugar beet root development to determine the cellular events that are temporally correlated with sucrose storage. The prestorage stage was characterized by a marked increase in root fresh weight and a low sucrose to glucose ratio. Carbon derived from ¹⁴C-sucrose accumulation was partitioned into protein and structural carbohydrate fractions and their amino acid, organic acid, and hexose precursors. The immature root contained high soluble acid invertase activity (V_max 20 micromoles per hour per milligram protein; K_m 2 to 3 millimolar) which disappeared prior to sucrose storage. Sucrose storage was characterized by carbon derived from ¹⁴C-sucrose uptake being partitioned into the sucrose fraction with little evidence of further metabolism. The onset of storage was accompanied by the appearance of sucrose synthetase activity (V_max 12 micromoles per hour per milligram protein; K_m 7 millimolar). Neither sucrose phosphate synthetase nor alkaline invertase activities were detected during beet development. Intact sugar beet plants (containing a 100-gram beet) exported 70% of the translocate to the beet, greater than 90% of which was retained as sucrose with little subsequent conversions.     

Inhibition by ethylene of polyamine biosynthetic enzymes enhanced lysine decarboxylase activity and cadaverine accumulation in pea seedlings

Exposing etiolated pea seedlings to ethylene which inhibited the activity of arginine decarboxylase and S-adenosylmethionine decarboxylase caused an increase in the level of cadaverine. The elevated level of cadaverine resulted from an increase in lysine decarboxylase activity in the tissue exposed to ethylene. The hormone did not affect the apparent K_m of the enzyme, but the apparent V_max was increased by 96%. While lysine decarboxylase activity in the ethylene-treated plants increased in both the meristematic and the elongation zone tissue, cadaverine accumulation was observed in the latter only. The enhancement by ethylene of the enzyme activity was reversed completely 24 hours after transferring the plants to an ethylene-free atmosphere. It is postulated that the increase in lysine decarboxylase activity, and the consequent accumulation of cadaverine in ethylene-treated plants, is of a compensatory nature as a response to the inhibition of arginine and S-adenosylmethionine decarboxylase activity provoked by ethylene.     

Effect of retinol deficiency and curcumin or turmeric feeding on brain Na+−K+ adenosine triphosphatase activity

The effect of retinol deficiency and curcumin and turmeric feeding on brain microsomal Na⁺-K⁺ ATPase activity was investigated. The brain Na⁺–K⁺ ATPase activity registered an increase of 148.5% as compared to the control group. Upon treating retinol deficient rats with curcumin or turmeric, the abnormally elevated activity showed a decrease of 36.9 and 47.1%, respectively, when compared to the retinol deficient group. An increase in V_max by 67% and K_m by 66% for ATP was observed in the retinol deficient group. Curcumin or turmeric fed retinol-deficient groups reduced the V_max by 25 and 33%, while K_m was reduced by 25 and 31%, respectively, compared to the retinol deficient group. Arrhenius plot of Na⁺–K⁺ ATPase showed a typical bi-phasic pattern in all the groups. Cholesterol: Phospholipid ratio showed a decrease in the retinol-deficient group by 67.8%, which showed a marked increase in curcumin or turmeric treated groups. Detergents could increase the Na⁺–K⁺ ATPase activity more in the control group than in the retinol deficient groups. Curcumin or turmeric improved the detergent action on the enzyme. Subsequent freezing and thawing over a period of 30 min decreased the enzyme activity by 22.8% in the retinol deficient group compared to 15.9% decrease in the control group. Curcumin or turmeric treated groups showed a decrease in the enzyme activity by 22.0 and 19.2%, respectively, when compared to the zero time in each group. In the presence of concanavalin-A (Con-A) there was only 52.4% stimulation in the enzyme activity in retinol deficient groups, compared to 108.0% in the control group. Curcumin or turmeric treated retinol-deficient groups showed a stimulation in the presence of con-A by 70 and 99.5%, respectively.     

Quantitative determination of calcium-activated myosin adenosine triphosphatase activity in rat skeletal muscle fibres

Summary A quantitative histochemical technique was developed for determining the kinetics of the calcium-activated myosin ATPase (Ca²⁺-myosin ATPase) reaction in rat skeletal muscle fibres. Using this technique, the maximum velocity (V_max) and the apparent Michaelis-Menten rate constant for ATP (K_app) of the Ca²⁺-myosin ATPase reaction were measured in type-identified fibres of the rat medial gastrocnemius (MG) muscle. The V_max and the K_app of the Ca²⁺-myosin ATPase reaction were lowest in type I fibres and highest (i.e., approx. two times greater) in type IIb fibres. The K_app in type IIa fibres was similar to that in type I. However, the V_max was 1.5 times greater in type IIa fibres, compared to type I fibres. Evidence is presented to suggest that the type IIb fibre population in the MG does not represent a single myosin isozyme. In addition, the broad range of V_max and K_app values indicates that there is marked heterogeneity in the myosin heavy chain and myosin light chain composition of myosin isozymes among individual fibres.     

Kinetic Analyses of the Substrate Inhibition of <Emphasis Type="Italic">Paramecium</Emphasis> Arginine Kinase

Paramecium tetraurelia expresses four types of arginine kinase (AK1–AK4). In a previous study, we showed that AK3 is characterized by typical arginine substrate inhibition, where enzymatic activity markedly decreases near a concentration of 1 mM of arginine substrate. This is in sharp contrast to the three other AK types, which obey the Michaelis–Menten reaction curve. Since cellular arginine concentration in another ciliate Tetrahymena is estimated to be 3–15 mM in vivo, Paramecium AK3 likely functions in conditions that are strongly affected by substrate inhibition. The purpose of this work is to find some novel aspect on the kinetic mechanism of the substrate inhibition of Paramecium AK3 enzyme. Substrate inhibition kinetics for AK3 were analyzed using three models and their validity were evaluated with three static parameters (R², AICc, and Sy.x). The most accurate model indicated that not only ES but also the SES complex reacts to form products, the latter being the complex with two substrates in the active center. The maximum reaction rate for the SES complex, V_max^SES?=?30.4 µmol Pi/min/mg protein, was one-eighth of the ES complex, V_max^ES?=?241.7. The dissociation constant for the SES complex (K_i^SES: 0.34 mM) was two times smaller than that of the ES complex (K_s^ES: 0.61 mM), suggesting that after the primary binding of the arginine substrate (ES complex formation), the binding of a second arginine to the secondarily induced inhibitory site is accelerated to form an SES complex with a lower V_max^SES. The same kinetics were used for the S79A, S80A, and V81A mutants. The results indicate that the S79 residue is significantly involved in the process of binding the second arginine substrate. Herein, the K_i^SES value was ten times (3.62 mM) the value for the wild-type (0.34 mM), weakening substrate inhibition. In contrast, V_max^ES and V_max^SES values for the mutants decreased by one-third, except for the V_max^SES of the S79A mutant, which had a value that was comparable with the value for the wild-type.     

Effect of pH on the Kinetic Parameters of NADP-Malic Enzyme from a C(4)Flaveria (Asteraceae) Species

We have used the pH variation in the kinetic parameters with respect to malate of NADP-malic enzyme purified from the C₄ species, Flaveria trinervia, to compare the pK values of its functional groups with those for the pigeon liver NADP-malic enzyme (MI Schimerlik, WW Cleland [1977] Biochemistry 16: 576-583) and the plant NAD-malic enzyme (KO Willeford, RT Wedding [1987] Plant Physiol 84: 1084-1087). Like the other enzymes, the C₄ enzyme has a group with a pK of about 6.0 (6.6 for the C₄ enzyme), as indicated from plots of the log V_max/K_m (V_max = maximum rate of catalysis) versus pH, which must lose a proton for malate binding and subsequent catalysis. The optimum ionization for the C₄ enzyme-NADP-Mg²⁺ complex occurs at pH 7.1 to 7.5. From pH 7.5 to 8.4, the K_m increases, but V_max remains constant. The log V_max/K_m plot in this pH range indicates a group with a pK of about 7.7. The other malic enzymes exhibit a similar pK. Above pH 8.4, deprotonation leads to a marked increase in K_m and a decrease in V_max for the C₄ enzyme. As in the case of the animal enzyme, the log V_max/K_m plot for the C₄ enzyme appears to approach a slope of two. The curve suggests an average pK of 8.4 for the groups involved, while the animal enzyme exhibits an average pK of 9.0. The NAD-malic enzyme does not exhibit any pK values at these high pK values. We hypothesize that the putative groups with the high pK values may be at least partially responsible for the ability of the C₄ NADP-malic enzyme to maintain high activity at pH 8.0 in illuminated chloroplasts.     

Kinetic study of the cellobiase activity of Trichoderma reesei cellulase complex at high substrate concentrations

At high cellobiose concentrations, the cellobiase activity of a Trichoderma reesei cellulase preparation does not follow Michaelis–Menten kinetics and shows substrate inhibition. Several rate equations were fitted to the initial rate-cellobiose concentration data. The best fit is obtained for a rate equation corresponding to partial substrate inhibition of cellobiase. In this case, the K_m, V_max and K_I values obtained are 1.1 mM, 16 IU ml^–1 and 26 mM, respectively.     

The use and misuse of V c,max in Earth System Models

Earth System Models (ESMs) aim to project global change. Central to this aim is the need to accurately model global carbon fluxes. Photosynthetic carbon dioxide assimilation by the terrestrial biosphere is the largest of these fluxes, and in many ESMs is represented by the Farquhar, von Caemmerer and Berry (FvCB) model of photosynthesis. The maximum rate of carboxylation by the enzyme Rubisco, commonly termed V _c,max, is a key parameter in the FvCB model. This study investigated the derivation of the values of V _c,max used to represent different plant functional types (PFTs) in ESMs. Four methods for estimating V _c,max were identified; (1) an empirical or (2) mechanistic relationship was used to relate V _c,max to leaf N content, (3) V _c,max was estimated using an approach based on the optimization of photosynthesis and respiration or (4) calibration of a user-defined V _c,max to obtain a target model output. Despite representing the same PFTs, the land model components of ESMs were parameterized with a wide range of values for V _c,max (?46 to +77 % of the PFT mean). In many cases, parameterization was based on limited data sets and poorly defined coefficients that were used to adjust model parameters and set PFT-specific values for V _c,max. Examination of the models that linked leaf N mechanistically to V _c,max identified potential changes to fixed parameters that collectively would decrease V _c,max by 31 % in C₃ plants and 11 % in C₄ plants. Plant trait data bases are now available that offer an excellent opportunity for models to update PFT-specific parameters used to estimate V _c,max. However, data for parameterizing some PFTs, particularly those in the Tropics and the Arctic are either highly variable or largely absent.