SASDCX3: Proline utilization A from Desulfovibrio vulgaris 3.0 mg/mL

基本情報

• 登録情報: データベース: SASBDB / ID: SASDCX3

試料

Proline utilization A from Desulfovibrio vulgaris 3.0 mg/mL

• Bifunctional protein PutA (protein), DvPutA, Desulfovibrio vulgaris (strain RCH1)

機能・相同性

分子機能:

proline dehydrogenase / proline dehydrogenase activity / oxidoreductase activity, acting on the aldehyde or oxo group of donors, NAD or NADP as acceptor / L-glutamate gamma-semialdehyde dehydrogenase / 1-pyrroline-5-carboxylate dehydrogenase activity / proline catabolic process to glutamate / proline biosynthetic process / DNA-binding transcription factor activity / DNA binding

ドメイン・相同性:

Proline utilization A, N-terminal / Proline utilization A N-terminal domain / 1-pyrroline-5-carboxylate dehydrogenase / Bifunctional protein PutA / Proline dehydrogenase domain / Proline dehydrogenase / FAD-linked oxidoreductase-like / Aldehyde dehydrogenase, glutamic acid active site / Aldehyde dehydrogenases glutamic acid active site. / Aldehyde dehydrogenase, cysteine active site / Aldehyde dehydrogenases cysteine active site. / Aldehyde dehydrogenase domain / Aldehyde dehydrogenase family / Aldehyde dehydrogenase, N-terminal / Aldehyde dehydrogenase, C-terminal / Aldehyde/histidinol dehydrogenase

構成要素:

Bifunctional protein PutA

• 生物種: Desulfovibrio vulgaris (strain RCH1) (バクテリア)
• 引用: ジャーナル: FEBS J / 年: 2017; タイトル: Biophysical investigation of type A PutAs reveals a conserved core oligomeric structure.; 著者: David A Korasick / Harkewal Singh / Travis A Pemberton / Min Luo / Richa Dhatwalia / John J Tanner / ; 要旨: Many enzymes form homooligomers, yet the functional significance of self-association is seldom obvious. Herein, we examine the connection between oligomerization and catalytic function for proline utilization A (PutA) enzymes. PutAs are bifunctional enzymes that catalyze both reactions of proline catabolism. Type A PutAs are the smallest members of the family, possessing a minimal domain architecture consisting of N-terminal proline dehydrogenase and C-terminal l-glutamate-γ-semialdehyde dehydrogenase modules. Type A PutAs form domain-swapped dimers, and in one case (Bradyrhizobium japonicum PutA), two of the dimers assemble into a ring-shaped tetramer. Whereas the dimer has a clear role in substrate channeling, the functional significance of the tetramer is unknown. To address this question, we performed structural studies of four-type A PutAs from two clades of the PutA tree. The crystal structure of Bdellovibrio bacteriovorus PutA covalently inactivated by N-propargylglycine revealed a fold and substrate-channeling tunnel similar to other PutAs. Small-angle X-ray scattering (SAXS) and analytical ultracentrifugation indicated that Bdellovibrio PutA is dimeric in solution, in contrast to the prediction from crystal packing of a stable tetrameric assembly. SAXS studies of two other type A PutAs from separate clades also suggested that the dimer predominates in solution. To assess whether the tetramer of B. japonicum PutA is necessary for catalytic function, a hot spot disruption mutant that cleanly produces dimeric protein was generated. The dimeric variant exhibited kinetic parameters similar to the wild-type enzyme. These results implicate the domain-swapped dimer as the core structural and functional unit of type A PutAs.; ENZYMES: Proline dehydrogenase (EC 1.5.5.2); l-glutamate-γ-semialdehyde dehydrogenase (EC 1.2.1.88).; DATABASES: The atomic coordinates and structure factor amplitudes have been deposited in the Protein Data Bank under accession number 5UR2. The SAXS data have been deposited in the SASBDB under the following accession codes: SASDCP3 (BbPutA), SASDCQ3 (DvPutA 1.5 mg·mL ), SASDCX3 (DvPutA 3.0 mg·mL ), SASDCY3 (DvPutA 4.5 mg·mL ), SASDCR3 (LpPutA 3.0 mg·mL ), SASDCV3 (LpPutA 5.0 mg·mL ), SASDCW3 (LpPutA 8.0 mg·mL ), SASDCS3 (BjPutA 2.3 mg·mL ), SASDCT3 (BjPutA 4.7 mg·mL ), SASDCU3 (BjPutA 7.0 mg·mL ), SASDCZ3 (R51E 2.3 mg·mL ), SASDC24 (R51E 4.7 mg·mL ), SASDC34 (R51E 7.0 mg·mL ).

登録者

• John Tanner (Mizzou, University of Missouri-Columbia, Columbia, MO, USA)

モデル

• モデル #1431: ; タイプ: dummy / ダミー原子の半径: 3.40 A / 対称性: P2 / カイ2乗値: 1.040; Omokage検索でこの集合体の類似形状データを探す (詳細)
• モデル #1432: ; タイプ: dummy / ソフトウェア: (5.0 r6293) / ダミー原子の半径: 4.25 A / カイ2乗値: 1.040; Omokage検索でこの集合体の類似形状データを探す (詳細)

試料

• 試料: 名称: Proline utilization A from Desulfovibrio vulgaris 3.0 mg/mL; 試料濃度: 3 mg/ml
• バッファ: 名称: 50 mM Tris-HCl, 50 mM NaCl, 0.5 mM EDTA, and 0.5 mM THP at pH 7.5.; pH: 7.5

要素 #679

名称: DvPutA / タイプ: protein / 記述: Bifunctional protein PutA / 分子量: 114.496 / 分子数: 2 / 由来: Desulfovibrio vulgaris (strain RCH1) / 参照: UniProt: A0A0E0T6R2
配列:

MHHHHHHSSG VDLGTENLYF QSMDQQHLDG KVVERGKEFF RSISGEAPSI FNKGWWTGKV MDWAMQNEDF KVQLFRFVDV LPYLNTSESL LRHIREYFAT EDADIPPVLK WGAGKAGIGG ALTAKLMGMT IRSNIEGMAR QFIIGDNSKE AVKGLAKLRK DGFTFTVDLL GEATVSEEES EAYAQGYHEV VDAIAREQEK WKALPGNGPV EGFDWGATPK VNVSIKPSAL YSQAKPVDVE GSVRGILSRL VPIYRKVVAM GGFLCIDMEQ LKYKEMTLEL FKRLRSDPEF RHYPHLSIVL QAYLRDTEKD LDDLLHWARS EKLPIGIRLV KGAYWDYETV IAKQNGWEIP VWTDKPESDI AYEKLAHRIL ENSDIVYFAC ASHNVRTIAA VMETALALNV PEHRYEFQVL YGMAEPVRKG LKNVAGRVRL YCPYGELIPG MAYLVRRLLE NTANESFLRQ SFAEGAALER LLENPQKTLH RLLAARPEPR AVEPGPGGLP PFTNDAMIDF TVPDNRKAFV EALADVRSRF GQTVPLYIGG RDVTTADLIP TTNPAKPAEV VASICQAGRP EIDDAIAAAK KAALTWRDTS PADRAAYLRR AADICRKRIW ELSAWQVVEV GKQWDQAYHD VTEGIDFLEY YAREMLRLGA PRRMGRAPGE HNHLFYQPKG IAAVIAPWNF PFAIAIGMAS AAIVTGNPVI FKPSSISSRI GYNLAEVFRE AGLPEGVFNY CPGRSSIMGD YLVEHPDISL ICFTGSMEVG LRIQEKAAKV QPGQRQCKRV IAEMGGKNAT IIDDDADLDE AVLQVLYSAF GFQGQKCSAC SRVIVLDAIY DRFIERLVKA ASSIHIGPSE DPSNYMGPVA DATLQKNVSD YIRIAEEEGR VLLKRTDLPA EGCYVPLTIV GDIRPEHRIA QEEIFGPVLA VMRAATFDEA LSIANGTRFA LTGAVFSRSP EHLDKARREF RVGNLYLNKG STGALVERQP FGGFAMSGVG SKTGGPDYLL QFMDPRVVTE NTMRRGFTPI DEDDDWIV

実験情報

• ビーム: 設備名称: Advanced Light Source (ALS) 12.3.1 (SIBYLS) / 地域: Berkeley, CA / 国: USA / 線源: X-ray synchrotron
• 検出器: 名称: MAR 165 CCD

スキャン

タイトル: Proline utilization A from Desulfovibrio vulgaris 3.0 mg/mL
測定日: 2012年6月8日 / 単位: 1/A /

	Min	Max
Q	0.0112	0.3214

距離分布関数 P(R)

ソフトウェア P(R): GNOM 5.0 / ポイント数: 510 /

	Min	Max
Q	0.01125	0.321359
P(R) point	1	510
R	0	160

結果

カーブのタイプ: single_conc /

	実験値	Porod
分子量	224 kDa	-
体積	-	295 nm3

	P(R)	P(R) error	Guinier	Guinier error
前方散乱 I0	1625	6.3	1600.1	7.63
慣性半径, Rg	4.55 nm	0.02	4.37 nm	0.03

	Min	Max
D	-	16
Guinier point	1	30