Methods in Enzymology

METHODS IN ENZYMOLOGY

SIDNEY P. COLOWICK NATHAN O . KAPLAN

VOLUME I

PREFACE V

CONTRIBUTORS TO VOLUME I X i

OUTLINE OF ORGANIZATION, VOLUME I xV

OUTLINE OF VOLUMES II-IV xxi i

Section I. General Preparative Procedure s

1. Tissue Slice Technique K. A. C. ELLIOTT 3 2. Tissue Homogenates VAN R. POTTER 10 3. Fractionation of Cell Components of Anima l Tissues GEORGE H . HOGEBOOM 16 4. Preparation of Mitochondria from Plants BERNARD AXELROD 19 5. Preparation of Chloroplasts and Disinte- grated Chloroplasts PAUL R. GORHAM 22 6. Methods of Extraction of Enzymes from Animal Tissues ROBERT K. MORTON 25 7. Extraction of Enzymes from Microorgan- isms (Bacteria and Yeast) I. C. GuNsALUS 51 8. Extraction of Soluble Enzymes from Higher Plants ALVIN NASON 62 9. Disintegration of Bacteria and Small Par- C. S. FRENCH and 64 titles by High-Pressure Extrusion H. W. MILNE R 10. Protein Fractionation on the Basis of Solu- ARDA ALDEN GREEN and 67 bility in Aqueous Solutions of Salts and WALTER L. HUGHE S Organic Solvents 11. Separation of Proteins by Use of Adsorbents SIDNEY P. COLOWICK 90 12. The Partition Chromatography of Enzymes R. R. PORTER 98

13. Chromatography of Enzymes on Ion Ex- change Resins C. H. W. HIRs 113

14. Special Techniques for Bacterial Enzymes . Enrichment Culture and Adaptive En - zymes OSAMU HAYAISHI 126

15. Separation of Proteins from Nucleic Acids LEON HEPPEL 137

16. Preparation of Buffers for Use in Enzym e Studies G, GOMORI 138

Section II. Enzymes of Carbohydrate Metabolism

17. Amylases, a and ß PETER BERNFELD 14 9

18. Pectic Enzymes Z. I. KERTESZ 15 8 19. Mucopolysaccharidases ALBERT DORFMAN 16 6 20. Cellulase Preparation from Helix pomatia (Snails) GEORGE DE STEVENS 173 21 . Polysaccharide Synthesis from Disaccha- rides EDWARD J. HEHRE 178 22. Phosphorylases from Plants W. J. WHELAN 192 23 . Muscle Phosphorylase GERTY T. Cora, 200 BARBARA ILLINGWORTH, and PATRICIA J. KELLER

24. The PR Enzyme of Muscle PATRICIA J. KELLER and 206 GERTY T. Corn 25. Amylo-1,6-glucosidase GERTY T. Cora 21 1 26. Polysaccharide Phosphorylase, Liver EARL W. SUTHERLAND 21 5 27. Branching Enzyme from Liver JOSEPH LARNER 222 28. Disaccharide Phosphorylases M . DOUDOROFF 225 29 . Hexoside Hydrolases SHLOMO HESTRIN, 23 1 DAVID S. FEINGOLD , and MICHAEL SCHRAMM 30 . Methods for Measuring Transglycosylase Activity of Invertases J. S. D. BACON 258 31. Glucuronidases WILLIAM H. FISHMAN and 262 PETER BERNFELD

32. Yeast Hexokinase MARGARET R. MCDONALD 269 33 . Animal Tissue Hexokinases ROBERT K. CRANE and 277 ALBERTO SOLE

34. Fructokinase (Ketohexokinase) H. G. HERS 286

35 . Galactokinase and Galactowaldenase LUIS F. LELOIR and 29 0 RAfm E . TRUCC o 36. Phosphoglucomutase from Muscle VICTOR A . NAJJAR 29 4

37. Phosphohexoisomerases from Muscle MILTON W . SLEIN 29 9

38 . Phosphohexokinase KUO-HUANG LING, 30 6 WILLIAM L. BYRNE, and HENRY A. LARDY

39. Aldolase from Muscle JOHN FULLER TAYLOR 31 0 40. Aldolase from Yeast WALTER CHRISTIAN 31 5

41 . Fructose-l-Phosphate Aldolase from Liver FRANZ LEUTHARDT and 320 H. P. WoLF

42a. Glucose-6-phosphate Dehydrogenase A. KORNBERG and 32 3 B . L. HORECKE R b. 6-Phosphogluconic Dehydrogenase B . L. HORECKER and 323 P. Z . SMYRNIOTIS 43 . Glucose-6-phosphate and 6-phosphogluconi c Dehydrogenases from Leuconostoc mesenteroides R. D. DEMoss 328 44. Glucose Dehydrogenase from Liver HAROLD J. STRECKER 33 5 45. Glucose Aerodehydrogenase (Glucose Oxi- dase) RONALD BENTLEY 34 0

46. Hexose Phosphate and Hexose Reductase JOHN B. WOLFF and 34 6 NATHAN O. KAPLAN 47. Gluconokinase SEYMOUR S. COHEN 35 0 48. Synthesis of Glucose-1,6-diphosphate Luis F. LELOIR and 35 4 RAfiL E . TRUCCo

49. Yeast Pentokinase HENRY Z. SABLE 357

50. Phosphoribomutase from Muscle HANS KLENOW 36 1 51. Pentose Phosphate Isomerase BERNARD AXELROD 363 52. Pentose Isomerases SEYMOUR S. COHEN 366 53. Transketolase from Liver and Spinach B . L. HORECKER and 371 P. Z . SMYRNIOTIS 54. Crystalline Transketolase from Baker's G . DE LA HABA and 375 Yeast E . RACKER 55. Transaldolase B. L. HORECKER and 381 P. Z . SMYRNIOTIS 56 . Deoxyribose Phosphate Aldolase (DR- Aldolase) E. RACKER 384 57 . Triosephosphate Isomerase from Calf Muscle GERWIN BEISENHERZ 38 7 58. a-Glycerophosphate Dehydrogenase from GERWIN BEISENHERZ, 39 1 Rabbit Muscle THEODOR BLUCHER, and KARL-HEINZ GARBADE 59. Glycerol Dehydrogenase from Aerobactor aerogenes ROBERT MAIN BURTON 397

60 . Glyceraldehyde-3-phosphate Dehydrogen - ase from Muscle SIDNEY F . VELICK 401 61 . Glyceraldehyde-3-phosphate Dehydrogen - ase from Yeast EDWIN G . KREBS 407 62 . TPN Triosephosphate Dehydrogenase from Plant Tissue MARTIN GIBBS 411 63 . Phosphoglycerate Kinase from Brewer' s Yeast THEODOR BtICHER 415 64. Phosphoglyceric Acid Mutases PETER OESPER 423 65. Enolase from Brewer's Yeast THEODOR B$CHER 427 66 . Pyruvate Kinase from Muscle THEODOR BiicHER and 435 G. PFLEIDERER 67. Lactic Dehydrogenase of Muscle ARTHUR KORNBERG 441 68. Lactic Dehydrogenase from Yeast MALCOLM DIXON 444 69. Lactic Dehydrogenase of Heart Muscle J. B . NEILANDS 449 70. Glyoxalases E . RACKER 454 71. Plant Carboxylases THOMAS P. SINGER 460 72. Acetoin Formation in Bacteria ELLIOT JUNI 471 73 . Phosphoroclastic Split of Pyruvate, Yieldin g Formate (E. soli) HAROLD J. STRECKER 476

74 . Phosphoroclastic Split of Pyruvate, Yieldin g Hydrogen (Clostridium butylicum) H. J. KoEPSELL 479

75. Phosphate-Linked Pyruvic Acid Oxidase L. P. HAGER and 482 from Lactobacillus delbrückii FRITZ LIPMANN 76 . Coenzyme A-Linked Pyruvic Oxidase (Ani - mal) S. KORKES 486 77 . Coenzyme A-Linked Pyruvic Oxidase (Bac- terial) S. KORKES 490 78 . Liver Alcohol Dehydrogenase ROGER BONNICHSEN and 495 NORMAN G. BRIN K

79. Alcohol Dehydrogenase from Baker's Yeast E . RACKER 500 80. TPN-Alcohol Dehydrogenase from Leuco- nostoc mesenteroides R. D . DEMoss 504 81 . Potassium-Activated Yeast Aldehyde De- hydrogenase SIMON BLACK 508 82. TPN-Linked Aldehyde Dehydrogenase from Yeast J. EDWIN SEEGMILLER 511

83. Liver Aldehyde Dehydrogenase E. RACKER 51 4

84. Aldehyde . Dehydrogenase from Clostridium E. R . STADTMAN and 51 8 kluyveri ROBERT M . BURTON 85. Flavin-Linked Aldehyde Oxidase HENRY R . MAHLER 523 86. Glycolic Acid Oxidase and Glyoxylic Acid Reductase ISRAEL ZELITCH 528 87. Formic Dehydrogenase from Peas ALVIN NASON and 536 HENRY N. LITTL E 88. Formic Hydrogenlyase from Escherichia coli MAX BOVARNICK 539

Section III. Enzymes of Lipid Metabolism

89. Fatty Acid Oxidation in Mitochondria ALBERT L. LEHNINGER 545

90. Fatty Acid Oxidation in Higher Plants P . K. STUMPF 549 91. Butyrate Enzymes of Clostridium kluyveri H. A. BARKER 551 92. Butyryl Coenzyme A Dehydrogenase HENRY R. MAHLER 553 93. Crystalline Crotonase from Ox Liver JOSEPH R . STERN 559

94. ß-Ketoreductase FEODOR LYNEN and 566 OTTO WIELAND 95. Enzymes of Acetoacetate Formation and Breakdown JOSEPH R. STERN 573 96. Aceto-CoA-Kinase MARY ELLEN JONES and 585 FRITZ LIPMAN N

97. Acetate Kinase of Bacteria (Acetokinase) IRwIN A. RosE 59 1 98. Phosphotransacetylase from Clostridium kluyveri E. R . STADTMAN 596 99. Coenzyme A Transphorase from Clostridium H. A . BARKER, 599 kluyveri E. R. STADTMAN, and ARTHUR KORNBERG 100. Deacylases (Thiol Esterase) JOHN GERGELY 602 101. Acetylation of Amines with Pigeon Liver Enzyme HERBERT TABOR 608 102. Acetylation of D-Glucosamine by Pigeo n Liver Extracts MORRIS SOODAK 61 2 103. Amino Acid Acetylase of Clostridium kluy- I. LIEBERMAN and 61 6 veri H. A . BARKE R

104. Choline Acetylase DAVID NACHMANSOHN and 61 9 IRWIN B . WILSON

105. Acetoacetate Decarboxylase H. W. SEELEY 624 106. Lipases M . BIER 627 107. Acetylcholinesterase DAVID NACHMANSOHN and 642 IRWIN B . WILSON

108. Dialkylphosphofluoridase A. MAZUR 65 1

109. Liver Esterase ELMER STOTZ 657 110. Phospholipases OSAMU HAYAISHI 660 111. Enzymatic Phospholipid Synthesis : Phos- phatidic Acid ARTHUR KORNBERG 67 3 112. Choline Oxidase J. H. QUASTEL 67 4 113. Cholesterol Dehydrogenase from a Myco- bacterium THRESSA C . STADTMAN 67 8

Section IV. Enzymes of Citric Acid Cycl e

114. Crystalline Condensing Enzyme from Pig Heart SEVERO OCHOA 685

115. Aconitase from Pig Heart Muscle CHRISTIAN B . ANFINSEN, JR . 695 116. Isocitric Dehydrogenase System (TPN ) from Pig Heart SEVERO OCHOA 69 9

117. Isocitric Dehydrogenase of Yeast (TPN) ARTHUR KORNBERG 70 5 118. Isocitric Dehydrogenase of Yeast (DPN) ARTHUR KORNBERG 707 119. Diphosphopyridine Nucleotide Isocitric De- G. W. E. PLAUT and 71 0 hydrogenase from Animal Tissues S.-C. SUN G 120. a-Ketoglutaric Dehydrogenase System and Phosphorylating Enzyme from Hear t Muscle SEYMOUR KAUFMAN 714

121. Succinic Dehydrogenase WALTER D. BONNER, JR . 72 2

122. Fumarase VINCENT MASSEY 72 9

123. Malic Dehydrogenase from Pig Heart SEVERO OCHOA 73 5

124. "Malic" Enzyme SEVERO OCHOA 739 125. Oxalacetic Carboxylase of Micrococcus lyso- deikticus DENIS HERBERT 75 3 126. Oxalacetate Synthesizing Enzyme MERTON F. UTTER and 75 8 KIYOSHI KURAHASH I

AUTHOR INDEX 765

SUBJECT INDEX 785