Microorganisms and methods for production of 4-hydroxybutyrate, 1,4-butanediol and related compounds

An isolated nucleic acid molecule selected from:

• (a) a nucleic acid molecule encoding a variant amino acid sequence of a parent polypeptide set forth in any one of:
• SEQ ID NOS:
• 439-518, wherein said variant amino acid sequence comprises two or more variant amino acid positions selected from a group consisting of:
• 12L, 15P, 34K, 42D, 44L, 47T, 51R, 51K, 56G, 57A, 58C, 58L, 58H, 59G, 66F, 67S, 67A, 71A, 91R, 104R, 106T, 107V, 127I, 128N, 129A, 129G, 129Y, 129I, 131W, 132A, 133M, 133V, 135P, 136A, 139S, 139T, 142L, 156C, 158V, 159G, 164A, 167P, 172G, 178G, 184M, 187K, 188G, 200M, 204L, 204R, 208A, 208L, 211C, 215M, 215A, 217Q, 226R, 227L, 227I, 229A, 230V, 230R, 230Q, 231I, 233Q, 233V, 234G, 241S, 242S, 242D, 243R, 243N, 243E, 248T, 252S, 253F, 253L, 253V, 253A, 258R, 259T, 260S, 260T, 260C, 265I, 265C, 265W, 265V, 266S, 269L, 2731, 273T, 273S, 273M, 273N, 279T, 280S, 281L, 284P, 288L, 289K, 291V, 294V, 295F, 298L, 310L, 310P, 317A, 317T, 327K, 333A, 349W, 360E, 370L, 370H, 395P, 396H, 396K, 396W, 396V, 398F, 401M, 406H, 411W, 416G, 416S, 417A, 419S, 426G, 427L, 427F, 427V, 429L, 429M, 437P, 439H, 441S, 446V, 446I, 447P, 456A, 464T, 464S, 466N, 468N, 47V, 113N, 177Y, 193L, 260V, 294D, 353N, 355A, 383Y, 13L, 16P, 32K, 40D, 42L, 45T, 45V, 49R, 49K, 54G, 55A, 56C, 56L, 56H, 57G, 64F, 65S, 65A, 69A, 89R, 102R, 104T, 105V, 111N, 125I, 126N, 127A, 127G, 127Y, 129W, 130A, 131M, 131V, 133P, 134A, 137S, 137T, 140L, 154C, 156V, 157G, 162A, 165P, 170G, 175Y, 176G, 182M, 185K, 186G, 191L, 198M, 202L, 202R, 206A, 206L, 209C, 213M, 213A, 215Q, 224R, 225I, 227A, 228V, 228R, 228Q, 229I, 231Q, 231V, 232G, 239S, 240S, 240D, 241R, 241N, 241E, 246T, 250S, 251F, 251L, 251V, 251A, 256R, 257T, 258S, 258T, 263I, 263C, 263W, 263V, 264S, 267L, 271I, 271T, 271S, 271M, 271N, 277T, 278S, 279L, 282P, 286L, 287K, 289V, 292V, 292D, 293F, 296L, 308L, 308P, 316A, 316T, 326K, 332A, 348W, 352N, 354A, 359E, 369L, 369H, 382Y, 394P, 395H, 395K, 395W, 395V, 397F, 400M, 405H, 133R, 146R, 148T, 149V, 155N, 170N, 171A, 171G, 171Y, 171I, 173W, 174A, 175M, 175V, 177P, 178A, 181S, 181T, 184L, 198C, 200V, 201G, 209P, 214G, 219Y, 220G, 226M, 229K, 230G, 235L, 242M, 246R, 250A, 250L, 253C, 257M, 257A, 259Q, 268R, 269I, 271A, 272V, 272R, 272Q, 275Q, 275V, 276G, 283S, 284S, 284D, 285R, 285N, 285E, 290T, 294S, 295L, 295V, 295A, 300R, 301T, 302S, 302T, 302C, 302V, 307I, 307C, 307W, 307V, 308S, 311L, 315I, 315T, 315S, 315M, 315N, 321T, 322S, 323L, 326P, 330L, 331K, 333V, 336V, 336D, 337F, 340L, 352L, 352P, 359A, 359T, 379K, 385A, 401, 405N, 407A, 412E, 55C, 55H, 55L, 63F, 64S, 64A, 68A, 88R, 101R, 103T, 104V, 110N, 124I, 125N, 126A, 126G, 126Y, 126I, 128W, 130M, 130V, 132P, 133A, 136S, 136T, 139L, 153C, 155V, 156G, 160A, 169G, 174Y, 175G, 181M, 184K, 185G, 190L, 197M, 201L, 201R, 205A, 205L, 208C, 212M, 212A, 214Q, 223R, 224L, 224I, 226A, 227V, 227R, 227Q, 228I, 231G, 238S, 239D, 240R, 240N, 240E, 245T, 248S, 250F, 250V, 255R, 256T, 257S, 257C, 257V, 262I, 262C, 262W, 262V, 263S, 266L, 270I, 270T, 270S, 270M, 270N, 276T, 277S, 278L, 281P, 285L, 286K, 288V, 291D, 292F, 314A, 334K, 340A, 357W, 360N, 362A, 367E, 377L, 377H, 388Y, 402P, 403H, 403K, 403W, 403V, 405F, and 408M;
• wherein the variant amino acid sequence can convert 4-hydroxybutyryl-CoA (4HB-CoA) to 4-hydroxybutyraldehyde (4-HBAld);
• and
• (b) a nucleic acid molecule that is complementary to the nucleic acid molecule of (a);
• wherein other than the two or more variant amino acid positions, the variant amino acid sequence has at least 90% sequence identity to any one of the parent polypeptide amino acid sequences of SEQ ID NOS:
• 439-518;
• and
• wherein the variant amino acid sequence is not the sequence of the parent polypeptide or a naturally occurring amino acid sequence.