Kinetic properties of Serratia marcescens adenosine 5'-diphosphate glucose pyrophosphorylase

Abstract

The regulatory properties of partially purified adenosine 5'-diphosphate-(ADP) glucose pyrophosphorylase from two Serratia marcescens strains (ATCC 274 and ATCC 15365) have been studied. Slight or negligible activation by fructose-P2, pyridoxal-phosphate, or reduced nicotinamide adenine dinucleotide phosphate (NADPH) was observed. These compounds were previously shown to be potent activators of the ADPglucose pyrophosphorylases from the enterics, Salmonella typhimurium, Enterobacter aerogenes, Enterobacter cloacae, Citrobacter freundii, Escherichia aurescens, Shigella dysenteriae, and Escherichia coli. Phosphoenolpyruvate stimulated the rate of ADPglucose synthesis catalyzed by Serratia ADPglucose pyrophosphorylase about 1.5-to 2-fold but did not affect the So.5 values (concentration of substrate required for 50% maximal stimulation) of the substrates, a-glucose-i-phosphate, and adenosine 5'-triphosphate. Adenosine 5'-monophosphate (AMP), a potent inhibitor of the enteric ADPglucose pyrophosphorylase, is an effective inhibitor of the S. marcescens enzyme. ADP also inhibits but is not as effective as AMP. Activators of the enteric enzyme counteract the inhibition caused by AMP. This is in contrast to what is observed for the S. marcescens enzyme. Neither phosphoenolpyruvate, fructose-diphosphate, pyridoxal-phosphate, NADPH, 3-phosphoglycerate, fructose-6-phosphate, nor pyruvate effect the inhibition caused by AMP. The properties of the S. marcescens HY strain and Serratia liquefaciens ADPglucose pyrophosphorylase were found to be similar to the above two S. marcescens enzynies with respect to activation and inhibition. These observations provide another example where the properties of an enzyme found in the genus Serratia have been found to be different from the properties of the same enzyme present in the enteric genera Escherichia, Salmonella, Shigella, Citrobacter, and Enterobacter. 193 on July 16, 2020 by guest

Glucose-1-phosphate adenylyltransferase

“…<1, 11,18,19,25,41,, 118> [1, 2, 4, 9-12, 14, 16, 17, 24-28, 30-36, 38, 39, 41, 42, 51, 53-55, 57]) [1-57] P diphosphate + ADP-glucose <1-22, 24, 26-61, 82-110, 117> [3, 5-8, 13, 15, 18-23, 29, 32, 34-38, 41, 45, 46, 48, 52, 54-56] S GTP + a-d-glucose 1-phosphate <44> (<44> can somewhat replace ATP [18]) (Reversibility: ? <44> [34]) [18] P diphosphate + GDP-glucose Inhibitors 8-azaADP-glucose <41> (<41> photoinactivation, competitive inhibition, ADP-glucose and ATP protects [34]) [34] 8-azaATP <41> (<41> photoinactivation, competitive inhibition, 0.5 mol/mol enzyme subunit for complete inactivation, ADP-glucose and ATP protects [34]) [34] ADP <2- 22, 24, 26-47, 49, 51, 53, 54, 56-61, 115, 117> (<2-22, 24, 26-37, 40, 41, 43-47, 49, 51, 53, 54, 56, 58-61> allosteric inhibitor [22,32]; <47> kinetics [32]; <2- 22, 24, 26-37, 40, 41, 43-47, 49, 51, 53, 54, 56, 58-61> [5,32]; <19> 1 mM [10]; <27-29> most effective inhibitor [32]; <11> less effective than phosphate [3]; <34, 58> inhibition under activated and unactivated conditions [30]; <59> less effective than AMP [23,32]; <28> most effective, fructose 6-phosphate or fructose 1,6-bisphosphate partially reverses [13]; <38> phosphoenolpyruvate reverses [13]; <59> phosphoenolpyruvate, fructose bisphosphate, pyridoxal phosphate, NADPH, 3-phosphoglycerate, fructose 6phosphate or pyruvate does not reverse [23]; <59> glycolytic metabolites do not affect inhibition [32]; <44, 53, 56> not [32]; <2- 37, 40, 41, 43-47, 49, 51, 53, 54, 56, 58-61> not…”

“…; <28, 38, 39, 42, 57-61> no activation[13,22]; <4> recombinant enzyme is insensitive to 3-phosphoglycerate[53])[1-8, 10, 11, 13, 23-25, 27, 29, 32, 35, 37, 41, 43-45, 47, 48, 52, 53, 55, 56] 4-pyridoxic acid 5-phosphate <41> (<41> activation[32])[32] 6-phosphogluconate <58> (<58> slight activation[22])[22] ADP <4, 8, 11, 13, 18, 41> (<4, 8, 11, 13, 18, 41> activation, less effective than 3-phosphoglycerate [29]; <23, 25, 28, 38, 39, 42, 57-61> no activation [11, 13]) [29] AMP <4, 8, 11, 13, 18, 41, 115> (<4, 8, 11, 13, 18, 41> activation, less effective than 3-phosphoglycerate[29]; <115> activation at pH 7[49]; <23, 25, 28, 38, 39, 42, 57-61> no activation[11,13])[29,49] d-arabinitol 1,5-diphosphate <41> (<41> activation[32])[32] d-fructose 1,6-bisphosphate <2-47, 49, 51, 53, 54, 56-61, 114, 115, 117> (<11, 23, 25, 59> slight activation[3,4,11,23]; <11,13> activation of pyrophosphor-olysis[4][5][6];…”

“…22,32] NADPH <2-22, 24, 26-47, 49, 51, 53, 54, 56-61> (<19> activation, slight[10]; <19, 28, 36-42, 57-61> activation[10,13,30,22,32]; <39,42,57,61> one of the most effective activators[13]; <28,38> no activation[13]; <27-29,41> no ac-<2-47,49,51,53,54,56-61> activation[11,13,22,23,29,30,32]; <2-22,24,26-37,40,41,43-47,49,51,53,54,56,58-61> allosteric activator[22,32]; <34,58> influence on K m and pH-optimum[30]; <41> wild-type enzyme and mutant SG14[32]; <38,39,42,57,61> one of the most effective activators, less effective for Enterobacter hafniae[13]; <27-29> no activation[32])[11,13,22, 23, 29, 30, 32] pyruvate <30-33, 45-47, 49, 51, 53, 54, 56, 115> (<30-33,45-47,49,51,53,54,56> activation, kinetics[32]; <30,41> activation of chimeric enzyme EA contains the N-terminus of Escherichia coli enzyme and the C-terminus of Agrobacterium tumefaciens enzyme with higher affinity than activation of chimeric enzyme AE contains the N-terminus of Agrobacterium tumefaciens enzyme and the C-terminus of Escherichia coli enzyme[40]; <11,18,23,25,59> not[3,4,8,11,23])[32,40,49] …”