The metabolism of D-myo-inositol 1,4,5-trisphosphate and D-myo-inositol 1,3,4,5-tetrakisphosphate by porcine skeletal muscle.
In soluble and particulate extracts from muscle D-myo-inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] and D-myo-inositol 1,3,4,5-tetrakisphosphate [Ins(1,3,4,5)P4] are metabolised stepwise to inositol. Ins(1,4,5)P3 is rapidly dephosphorylated to D-myo-inositol 1,4-bisphosphate then to D-myo-inositol 4-phosphate and finally inositol. In soluble extracts Ins(1,3,4,5)P4 is dephosphorylated to D-myo-inositol 1,3,4-trisphosphate then sequentially to D-myo-inositol 3,4-bisphosphate, D-myo-inositol 3-phosphate and inositol, while in particulate extracts D-myo-inositol 1,3-bisphosphate is the predominant inositol bisphosphate formed. Dephosphorylation of these inositol polyphosphates is Mg2+ dependent and inhibited by D-2,3-bisphosphoglyceric acid. Ins(1,4,5)P3 is also phosphorylated to form Ins(1,3,4,5)P4 in soluble extracts by Ins(1,4,5)P3 3-kinase. Ins(1,4,5)P3 3-kinase activity is Mg2+ and ATP dependent and is stimulated by Ca2+ and calmodulin. Particulate (sarcotubular) inositol polyphosphate 5-phosphatase (5-phosphatase) is found in membranes which are intimately involved in excitation-contraction coupling and the generation of the primary Ca2+ signal of muscle cells. Particulate 5-phosphatase had the highest specific activity in the transverse-tubule membrane, when compared to the terminal cisternae and longitudinal-tubule membranes of the sarcoplasmic reticulum. Particulate Ins(1,3,4,5)P4-3-phosphatase activity was also detected after fractionation of solubilised sarcotubular membranes by DEAE-Sephacel. Particulate 5-phosphatase activity was purified 25,600-fold to a specific activity of 25.6 mumol Ins(1,4,5)P3 hydrolysed.min-1.mg protein-1, after DEAE-Sephacel and novel affinity chromatography using D-2,3-bisphosphoglycerate/agarose and Sepharose-4B-immobilised Ins(1,4,5)P3-analog matrices. Purified particulate 5-phosphatase had apparent Km of 46.3 microM and 1.9 microM and Vmax of 115 and 0.046 mumol substrate hydrolysed.min-1.mg protein-1, for Ins(1,4,5)P3 and Ins(1,3,4,5)P4, respectively. In contrast, purified soluble type I 5-phosphatase had apparent Km of 8.9 microM and 1.1 microM and Vmax of 3.55 and 0.13 mumol substrate hydrolysed.min-1.mg protein-1, for Ins(1,4,5P3 and Ins(1,3,4,5)P4, respectively. As in other cells, muscle 5-phosphatases have a lower affinity, but a higher capacity to metabolise Ins(1,4,5)P3 than Ins(1,3,4,5)P4. Soluble type I 5-phosphatase may have a functional role in the metabolism of both inositol polyphosphates, while particulate 5-phosphatase may primarily metabolise Ins(1,4,5)P3. Purified Ins(1,4,5)P3 3-kinase had an apparent Km of 0.42 microM and a Vmax of 4.12 nmol Ins(1,4,5)P3 phosphorylated.min-1.mg protein-1. The profile of inositol polyphosphate metabolism in muscle is similar to that reported in other tissues.(ABSTRACT TRUNCATED AT 400 WORDS)
Division of Biochemistry and Molecular Biology, John Curtin School of Medical Research, Australian National University, Canberra.
P S
PS
Foster
S P
SP
Hogan
P M
PM
Hansbro
R
R
O'Brien
B V
BV
Potter
S
S
Ozaki
M A
MA
Denborough
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002458:Cell Fractionation; D002851:Chromatography, High Pressure Liquid; D007294:Inositol / Q000378:metabolism; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism*; D007295:Inositol Phosphates / Q000378:metabolism*; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D009132:Muscles / Q000378:metabolism*; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D012995:Solubility; D013347:Subcellular Fractions / Q000378:metabolism; D013552:Swine
D007295:Inositol Phosphates; C081655:inositol-1,3,4,5-tetrakisphosphate; D007294:Inositol; D015544:Inositol 1,4,5-Trisphosphate; D000255:Adenosine Triphosphate; D017853:Phosphotransferases (Alcohol Group Acceptor); C051038:Inositol 1,4,5-trisphosphate 3-kinase; D008274:Magnesium; D002118:Calcium