SASDCP3: Proline utilization A from Bdellovibrio bacteriovorus

Basic information

• Entry: Database: SASBDB / ID: SASDCP3

Sample

Proline utilization A from Bdellovibrio bacteriovorus

• Bifunctional protein PutA (protein), Bdellovibrio bacteriovorus (strain ATCC 15356 / DSM 50701 / NCIB 9529 / HD100)

Function / homology

Function:

proline dehydrogenase / proline dehydrogenase activity / 1-pyrroline-5-carboxylate dehydrogenase activity / L-glutamate gamma-semialdehyde dehydrogenase / proline catabolic process to glutamate / proline biosynthetic process / DNA-binding transcription factor activity / DNA binding

Domain/homology:

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

Component:

Bifunctional protein PutA

• Biological species: Bdellovibrio bacteriovorus (strain ATCC 15356 / DSM 50701 / NCIB 9529 / HD100) (bacteria)
• Citation: Journal: FEBS J / Year: 2017; Title: Biophysical investigation of type A PutAs reveals a conserved core oligomeric structure.; Authors: David A Korasick / Harkewal Singh / Travis A Pemberton / Min Luo / Richa Dhatwalia / John J Tanner / ; Abstract: 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 ).

Contact author

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

Models

• Model #1419: ; Type: dummy / Radius of dummy atoms: 2.90 A / Symmetry: P2 / Chi-square value: 1.168; Search similar-shape structures of this assembly by Omokage search (details)
• Model #1420: ; Type: dummy / Software: (5.0 r6293) / Radius of dummy atoms: 4.25 A / Chi-square value: 1.168; Search similar-shape structures of this assembly by Omokage search (details)

Sample

• Sample: Name: Proline utilization A from Bdellovibrio bacteriovorus
• Buffer: Name: 50 mM Tris, 125 mM NaCl, 1 mM EDTA, and 1 mM tris(3-hydroxypropyl)phosphine (THP) at pH 7.5,; pH: 7.5

Entity #678

Type: protein / Description: Bifunctional protein PutA / Formula weight: 109.504 / Num. of mol.: 2
Source: Bdellovibrio bacteriovorus (strain ATCC 15356 / DSM 50701 / NCIB 9529 / HD100)
References: UniProt: Q6MNK1
Sequence:

GHMNDIQSQI VSRGEEILKR MESQSKASIF SKDFWYGSIM EWSMKNEKFK TNMFRFVDVL PSINSGDEVA RHLKEYFSED GGTLPPVFNV GLGLGSLAPG LMAGAIKKNV MGMAKMFITG ESPDEALPVL KKARKNKMTF TVDILGEATL SEKEAQDYSN KYMELVTWLA KDAEKWDEVP QIDRDHEGAL PKVNVSVKMT ALYSQIKDAA WDESKKILKD RLRPVFRLGM EKGVFVNLDM EQYSVKHLTL EVFTELINEP EFKNYKFFGI VIQAYLRDSF EDVKSLTEFA QKRGTPFWVR LVKGAYWDYE TIEAEQRGWP VPVYTNKAES DANYELCAKY LLENIKFIRP AFASHNVRTL AACMLYAEKL NIPKEALEFQ MLYGMAEPIK KTIVDMGYRM REYAPVGELI PGMAYLVRRL LENTSNESWL RGKFADNKSM AELLKDPAQG LTPTSPVIPK KPGKFYNEPL LDFAVKADRE KMLKALAEAK ASLPVNVNIV INNKELQSGK IFDRVNPSQS DQIVGKIQMA TTEQAEQAMQ AAQTAYKTWK NVPCEQRAAL VDKLADIMTR DRFKLIATQV LEVGKPWAEA DGDIGEAIDF CRYYARHMRE LQKPLRVGGL PGELSHYIYK SRGVTAVIAP WNFPLAILAG MVTAAAVAGN TVVMKPAEQS TVVAWGLMKM IQEAGFPQGV INFLPGYGEE VGEYIVNHKY TTTIAFTGSK AVGLHIMNRA AVVQPGQQHV KRCIIEMGGK NAVIIDNDAD LDEAVDGVIY SAFGFSGQKC SAASRVIVLD EVYDRFVDRL VETAKSIEIH PAENPKAYMG PVVDKEAYDR ILGTIAEAEK NHKLLFKGSV PGGGFFAPPT IFGDVPGDAK LAQAEIFGPV VAVIRAKNLD QALDIANSTE YALTGGVFSR SPANINRVKE ELEVGNLYVN RGITGAMVDR HPFGGFKMSG IGSKTGGPDY LKQYMEPACV TENTLRRGFA PAEE

Experimental information

• Beam: Instrument name: Advanced Light Source (ALS) 12.3.1 (SIBYLS); City: Berkeley, CA / 国: USA / Type of source: X-ray synchrotronSynchrotron
• Detector: Name: MAR 165 CCD

Scan

Title: Proline utilization A from Bdellovibrio bacteriovorus
Measurement date: Jun 8, 2012 / Unit: 1/A /

	Min	Max
Q	0.0112	0.3208

Distance distribution function P(R)

Sofotware P(R): GNOM 4.5a / Number of points: 509 /

	Min	Max
Q	0.01125	0.3208
P(R) point	1	509
R	0	140

Result

Type of curve: single_conc /

	Experimental	Porod
MW	225 kDa	-
Volume	-	287 nm3

	P(R)	P(R) error	Guinier	Guinier error
Forward scattering, I0	1114	3.1	1130.4	6.8
Radius of gyration, Rg	4.6 nm	0.01	4.49 nm	0.04

	Min	Max
D	-	14
Guinier point	1	30