ANTIBODY SHOP

Services
Name:
Phone:
E-mail:
Message

HydA | Iron-hydrogenase HydA1/HydA2

345 €

AS09 514  |  Clonality: Polyclonal  |  Host: Rabbit  |  Reactivity: Chlamydomonas reinhardtii

PRODUCT INFORMATION IN PDF

Qty: 
Availability:
14 st
Delivery:
Shipping:
Item No:
AS09 514

Info: Product suggestions Add review
product information
Background

Iron-hydrogenase HydA2 is catalyzing reversible oxidation of molecular hydrogen. In Chlamydomonas the protein is present in low levels of 1 µg/liter of culture.Synonyme: HYD1/1

Immunogen

Recombinant, full length Chlamydomonas reinhardtii HydA-2 Q8VZZ0

Host Rabbit
Clonality Polyclonal
Clone
Purity Serum
Format Lyophilized
Quantity 100 µl
Reconstitution For reconstitution add 100 µl of sterile water.
Storage

Store lyophilized/reconstituted at -20°C; once reconstituted make aliquots to avoid repeated freeze-thaw cycles. Please, remember to spin tubes briefly prior to opening them to avoid any losses that might occur from lyophilized material adhering to the cap or sides of the tubes.

Tested applications Western Blot (WB)
Related products

AS09 600 | anti-HydA | Iron-hydrogenase HydA2 rabbit antibodies

Collection of antibodies to Chlamydomonas reinhardtii

Algal protein extraction buffer

Secondary antibodies

Additional information

In Chlamydomonas HydA is present in low levels of 1 µg/liter of culture. Therefore, an induction of cells by anaerobic adaptation or sulfur depravation (10 x higher amount than with anaerobic adaptation) is necessary for successful detection using this antibody. Methods of HydA induction are described in Hemschemeier et al. 2009.

To detect HydA1/A2 in Chlamydomonas extracts amount loaded per well corresponds to 2 µg of chlorophyll for sulfur deprived cells, where relatively much HydA1 is synthesized or corresponds to 2-4 µg of artificially anaerobic induced cultures, where the HydA1 protein level is lower. This antibody is recognizing 1 ng of recombinant HydA protein.

application information
Recommended dilution

1 : 5000 with standard ECL (WB)

Expected | apparent MW

53.7 | 48 kDa (after transit peptide is cleaved)

Confirmed reactivity

Chlamydomonas reinhardtii

Predicted reactivity

Ostreococcus sp.

Not reactive in

No confirmed exceptions from predicted reactivity known in the moment

Additional information

HydA1 (497aa) has a calculated MW of 53.1 kDa, but this is including the signal peptide, which gets cleaved off. The protein without TP has a calculated MW of 47.5 kDa and runs according to its size at about 48 kDa.

HydA2 (505aa) has a calculated MW of 53.7 kDa, but this is including the signal peptide, which gets cleaved off. The protein without TP can only be estimated, since the cleavage site is known only from in silico analysis. It has a calculated MW of 47.3 kDa and should run in the gel also according to its size.

This antibody is binding recombinant HydA1/2 protein.

Selected references Wei et al. (2017). Light Intensity is Important for Hydrogen Production in NaHSO3-Treated Chlamydomonas reinhardtii. Plant Cell Physiol. 2017 Mar 1;58(3):451-457. doi: 10.1093/pcp/pcw216.
Eilenberg et al. (2016). The dual effect of a ferredoxin-hydrogenase fusion protein in vivo: successful divergence of the photosynthetic electron flux towards hydrogen production and elevated oxygen tolerance. Biotechnol Biofuels. 2016 Aug 30;9(1):182. doi: 10.1186/s13068-016-0601-3. eCollection 2016.
Liran et al. (2016). Microoxic Niches within the Thylakoid Stroma of Air-Grown Chlamydomonas reinhardtii Protect [FeFe]-Hydrogenase and Support Hydrogen Production under Fully Aerobic Environment. Plant Physiol. 2016 Sep;172(1):264-71. doi: 10.1104/pp.16.01063. Epub 2016 Jul 21.
Reifschneider-Wegner et al. (2014). Expression of the [FeFe] hydrogenase in the chloroplast of Chlamydomonas reinhardtii. Int J of Hydrogen Energy, Volume 39, Issue 8, 6 March 2014, Pages 3657–3665.
Pinto et al. (2013). Rubisco mutants of Chlamydomonas reinhardtii enhance photosynthetic hydrogen production. Appl Microbiol Biotechnol. May 7.
Magneschi et al. (2012). A Mutant in the ADH1 Gene of Chlamydomonas reinhardtii Elicits Metabolic 2 Restructuring during Anaerobiosis. Plant Physiol. January 23 (ahead of print).

Application example

50ng of purified protein (HydA1 and HydA2) were separated on 10% SDS-PAGE and blotted 25 min to PVDF membrane. Filters were blocked 1h with 3% low-fat milk powder in PBS-T (0.1% TWEEN 20) and probed with anti-HydA1/2 (AS09 514, 1:5000, over night at 4°C) and secondary anti-rabbit (1:10 000, 1 h) antibody (HRP conjugated, manufacture Pierce) in PBS-T containing 3% low fat milk powder. Antibody incubations were followed by washings in PBS-T (10, +10min and PBS (+5, +5 min). All washing steps were performed at RT with agitation. Signal was detected with ECL (Millipore) using CCD camer. Exposure time was 20 min.

The heterolog expressed proteins have both calculated MWs of 51 kDa (due to the tag) and run according to their size.

Courtesy Dr. Thomas Happe

 


western blot using anti-Chlamydomonas HydA antibodies

 


||| For other applications, usage on species other than stated above or any other questions, please use the LiveChat option or contact us at support@agrisera.com